ARAF, CPTAC-1339 - CPTAC Assay Portal (original) (raw)
Please include the following statement when referencing the CPTAC Assay Portal
We would like to acknowledge the National Cancer Institute’s Clinical Proteomic Tumor Analysis Consortium (CPTAC) Assay Portal (assays.cancer.gov) for developing assays and establishing criteria for the assays described in this publication.
Protein Sequence hover to view complete sequence
CPTAC-5775: View additional IGTGSFGTVFR data
non-CPTAC-5533: View additional IGTGSFGTVFR data
non-CPTAC-3958: View additional IGDFGLATVK data
| 10 | 20 | 30 | 40 | 50 |
|---|---|---|---|---|
| MEPPRGPPAN | GAEPSRAVGT | VKVYLPNKQR | TVVTVRDGMS | VYDSLDKALK |
| 60 | 70 | 80 | 90 | 100 |
| VRGLNQDCCV | VYRLIKGRKT | VTAWDTAIAP | LDGEELIVEV | LEDVPLTMHN |
| 110 | 120 | 130 | 140 | 150 |
| FVRKTFFSLA | FCDFCLKFLF | HGFRCQTCGY | KFHQHCSSKV | PTVCVDMSTN |
| 160 | 170 | 180 | 190 | 200 |
| RQQFYHSVQD | LSGGSRQHEA | PSNRPLNELL | TPQGPSPRTQ | HCDPEHFPFP |
| 210 | 220 | 230 | 240 | 250 |
| APANAPLQRI | RSTSTPNVHM | VSTTAPMDSN | LIQLTGQSFS | TDAAGSRGGS |
| 260 | 270 | 280 | 290 | 300 |
| DGTPRGSPSP | ASVSSGRKSP | HSKSPAEQRE | RKSLADDKKK | VKNLGYRDSG |
| 310 | 320 | 330 | 340 | 350 |
| YYWEVPPSEV | QLLKRIGTGS | FGTVFRGRWH | GDVAVKVLKV | SQPTAEQAQA |
| 360 | 370 | 380 | 390 | 400 |
| FKNEMQVLRK | TRHVNILLFM | GFMTRPGFAI | ITQWCEGSSL | YHHLHVADTR |
| 410 | 420 | 430 | 440 | 450 |
| FDMVQLIDVA | RQTAQGMDYL | HAKNIIHRDL | KSNNIFLHEG | LTVKIGDFGL |
| 460 | 470 | 480 | 490 | 500 |
| ATVKTRWSGA | QPLEQPSGSV | LWMAAEVIRM | QDPNPYSFQS | DVYAYGVVLY |
| 510 | 520 | 530 | 540 | 550 |
| ELMTGSLPYS | HIGCRDQIIF | MVGRGYLSPD | LSKISSNCPK | AMRRLLSDCL |
| 560 | 570 | 580 | 590 | 600 |
| KFQREERPLF | PQILATIELL | QRSLPKIERS | ASEPSLHRTQ | ADELPACLLS |
| 606 | ||||
| AARLVP |
Data source: UniProt
Position of Targeted Peptide Analytes Relative to SNPs, Isoforms, and PTMs
Uniprot Database Entry PhosphoSitePlus ®
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to view detailed assay information below
All other points link out to UniProt
Phosphorylation Acetylation Ubiquitylation Other
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Assay Details for CPTAC-1339 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
DS[+80.0]GYYWEVPPSEVQLLK
Modification Type
Phospho (ST)
Protein - Site of Modification
299
Peptide - Site of Modification
2
Peptide Start
298
Peptide End
314
CPTAC ID
CPTAC-1339
Peptide Molecular Mass
2,088.9445
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
Digested Cell Lysate
Submitting Laboratory
Broad Institute
Submitting Lab PI
Steven A. Carr
Publication
View Details (opens in a new window)
Abelin, et al. Mol Cell Proteomics. 2016 May;15(5):1622-41. doi: 10.1074/mcp.M116.058354. Epub 2016 Feb 24. PMID:26912667
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
Quantiva TSQ
Internal Standard
medium stable isotope peptide (delta 6 amu)
Peptide Standard Purity
Crude (~60%)
Peptide Standard Label Type
13C at C-terminus K
LC
Easy NanoLC1000
Column Packing
Reprosil C18, 3 um, 200A
Column Dimensions
0.075 x 100 mm
Flow Rate
300 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV
| n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y9 (1+) | 23.8 | 4.1 | 4.1 | 26.7 | 6.5 | 8.7 | 35.8 | 7.7 | 9.6 | 15 | 15 | 15 |
| b8 (1+) | 26.9 | 10.6 | 8.2 | 33.8 | 10 | 17.2 | 43.2 | 14.6 | 19.1 | 15 | 15 | 15 |
| b8-98 (1+) | 23.5 | 9.6 | 7.5 | 29 | 13.3 | 14.9 | 37.3 | 16.4 | 16.7 | 15 | 15 | 15 |
| sum | 14.7 | 4 | 4.2 | 14.8 | 4.9 | 9.7 | 20.9 | 6.3 | 10.6 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for CPTAC-1343 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
QHEAPSNRPLNELLT[+80.0]PQGPS[+80.0]PR
Modification Type
Phospho (ST), Phospho (ST)
Protein - Site of Modification
181, 186
Peptide - Site of Modification
15, 20
Peptide Start
167
Peptide End
188
CPTAC ID
CPTAC-1343
Peptide Molecular Mass
2,597.1789
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
digested cell lysate
Submitting Laboratory
Broad Institute
Submitting Lab PI
Steven A. Carr
Publication
View Details (opens in a new window)
Mol Cell Proteomics. 2016 May;15(5):1622-41. doi: 10.1074/mcp.M116.058354. Epub 2016 Feb 24. Reduced-representation Phosphosignatures Measured by Quantitative Targeted MS Capture Cellular States and Enable Large-scale Comparison of Drug-induced Phenotypes. Abelin JG1, Patel J1, Lu X1, Feeney CM1, Fagbami L1, Creech AL1, Hu R1, Lam D1, Davison D1, Pino L1, Qiao JW1, Kuhn E1, Officer A1, Li J2, Abbatiello S1, Subramanian A1, Sidman R2, Snyder E3, Carr SA1, Jaffe JD4.
View Details (opens in a new window)
Mol Cell Proteomics. 2014 Jul;13(7):1690-704. doi: 10.1074/mcp.M113.036392. Epub 2014 Apr 9. Ischemia in tumors induces early and sustained phosphorylation changes in stress kinase pathways but does not affect global protein levels. Mertins P1, Yang F2, Liu T2, Mani DR3, Petyuk VA2, Gillette MA3, Clauser KR3, Qiao JW3, Gritsenko MA2, Moore RJ2, Levine DA4, Townsend R5, Erdmann-Gilmore P5, Snider JE5, Davies SR5, Ruggles KV6, Fenyo D6, Kitchens RT5, Li S5, Olvera N4, Dao F4, Rodriguez H7, Chan DW8, Liebler D9, White F10, Rodland KD2, Mills GB11, Smith RD2, Paulovich AG12, Ellis M5, Carr SA1
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
Quantiva
Internal Standard
light stable isotope peptide
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N at C-terminus R
LC
Easy NanoLC1000
Column Packing
Reprosil C18, 1.9um, 200A
Column Dimensions
0.075 x 150 mm
Flow Rate
200 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y18 (2+) | 16.1 | 11.7 | 9.7 | 15.6 | 13.6 | 19.6 | 22.4 | 17.9 | 21.9 | 13 | 13 | 15 |
| y7-98 (1+) | 8.4 | 7.1 | 4.4 | 9.2 | 7.7 | 14.1 | 12.5 | 10.5 | 14.8 | 15 | 15 | 15 |
| y7 (1+) | 6.5 | 5.3 | 6.4 | 7 | 6.2 | 10.2 | 9.6 | 8.2 | 12 | 15 | 15 | 15 |
| sum | 4.1 | 5.6 | 4.2 | 5.1 | 5.7 | 12.8 | 6.5 | 8 | 13.5 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for CPTAC-1344 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
QHEAPSNRPLNELLT[+80.0]PQGPSPR
Modification Type
Phospho (ST)
Protein - Site of Modification
181
Peptide - Site of Modification
15
Peptide Start
167
Peptide End
188
CPTAC ID
CPTAC-1344
Peptide Molecular Mass
2,517.2126
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
digested cell lysate
Submitting Laboratory
Broad Institute
Submitting Lab PI
Steven A. Carr
Publication
View Details (opens in a new window)
Mol Cell Proteomics. 2016 May;15(5):1622-41. doi: 10.1074/mcp.M116.058354. Epub 2016 Feb 24. Reduced-representation Phosphosignatures Measured by Quantitative Targeted MS Capture Cellular States and Enable Large-scale Comparison of Drug-induced Phenotypes. Abelin JG1, Patel J1, Lu X1, Feeney CM1, Fagbami L1, Creech AL1, Hu R1, Lam D1, Davison D1, Pino L1, Qiao JW1, Kuhn E1, Officer A1, Li J2, Abbatiello S1, Subramanian A1, Sidman R2, Snyder E3, Carr SA1, Jaffe JD4.
View Details (opens in a new window)
Mol Cell Proteomics. 2014 Jul;13(7):1690-704. doi: 10.1074/mcp.M113.036392. Epub 2014 Apr 9. Ischemia in tumors induces early and sustained phosphorylation changes in stress kinase pathways but does not affect global protein levels. Mertins P1, Yang F2, Liu T2, Mani DR3, Petyuk VA2, Gillette MA3, Clauser KR3, Qiao JW3, Gritsenko MA2, Moore RJ2, Levine DA4, Townsend R5, Erdmann-Gilmore P5, Snider JE5, Davies SR5, Ruggles KV6, Fenyo D6, Kitchens RT5, Li S5, Olvera N4, Dao F4, Rodriguez H7, Chan DW8, Liebler D9, White F10, Rodland KD2, Mills GB11, Smith RD2, Paulovich AG12, Ellis M5, Carr SA1
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
Quantiva
Internal Standard
light stable isotope peptide
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N at C-terminus R
LC
Easy NanoLC1000
Column Packing
Reprosil C18, 1.9um, 200A
Column Dimensions
0.075 x 150 mm
Flow Rate
200 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y18 (2+) | 16.3 | 9.5 | 7.1 | 19.3 | 10.8 | 9.8 | 25.3 | 14.4 | 12.1 | 14 | 15 | 15 |
| y7 (1+) | 5.7 | 6.3 | 7.7 | 8.2 | 15.3 | 11.3 | 10 | 16.5 | 13.7 | 15 | 15 | 15 |
| y8-98 (1+) | 13.6 | 13.9 | 8.9 | 15.9 | 23.2 | 12.5 | 20.9 | 27 | 15.3 | 15 | 15 | 15 |
| sum | 5.8 | 6.4 | 5.5 | 8.1 | 11.8 | 9.1 | 10 | 13.4 | 10.6 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for CPTAC-1345 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
QHEAPSNRPLNELLTPQGPS[+80.0]PR
Modification Type
Phospho (ST)
Protein - Site of Modification
186
Peptide - Site of Modification
20
Peptide Start
167
Peptide End
188
CPTAC ID
CPTAC-1345
Peptide Molecular Mass
2,517.2126
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
digested cell lysate
Submitting Laboratory
Broad Institute
Submitting Lab PI
Steven A. Carr
Publication
View Details (opens in a new window)
Mol Cell Proteomics. 2016 May;15(5):1622-41. doi: 10.1074/mcp.M116.058354. Epub 2016 Feb 24. Reduced-representation Phosphosignatures Measured by Quantitative Targeted MS Capture Cellular States and Enable Large-scale Comparison of Drug-induced Phenotypes. Abelin JG1, Patel J1, Lu X1, Feeney CM1, Fagbami L1, Creech AL1, Hu R1, Lam D1, Davison D1, Pino L1, Qiao JW1, Kuhn E1, Officer A1, Li J2, Abbatiello S1, Subramanian A1, Sidman R2, Snyder E3, Carr SA1, Jaffe JD4.
View Details (opens in a new window)
Mol Cell Proteomics. 2014 Jul;13(7):1690-704. doi: 10.1074/mcp.M113.036392. Epub 2014 Apr 9. Ischemia in tumors induces early and sustained phosphorylation changes in stress kinase pathways but does not affect global protein levels. Mertins P1, Yang F2, Liu T2, Mani DR3, Petyuk VA2, Gillette MA3, Clauser KR3, Qiao JW3, Gritsenko MA2, Moore RJ2, Levine DA4, Townsend R5, Erdmann-Gilmore P5, Snider JE5, Davies SR5, Ruggles KV6, Fenyo D6, Kitchens RT5, Li S5, Olvera N4, Dao F4, Rodriguez H7, Chan DW8, Liebler D9, White F10, Rodland KD2, Mills GB11, Smith RD2, Paulovich AG12, Ellis M5, Carr SA1
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
Quantiva
Internal Standard
light stable isotope peptide
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N at C-terminus R
LC
Easy NanoLC1000
Column Packing
Reprosil C18, 1.9um, 200A
Column Dimensions
0.075 x 150 mm
Flow Rate
200 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y18-98 (2+) | 10 | 10.9 | 11.7 | 8.8 | 11.6 | 11.2 | 13.3 | 15.9 | 16.2 | 15 | 15 | 15 |
| y18 (2+) | 7.3 | 7.2 | 6.3 | 9.2 | 9 | 9.5 | 11.7 | 11.5 | 11.4 | 15 | 15 | 15 |
| y7 (1+) | 6 | 6.8 | 10.5 | 8.1 | 15.4 | 10 | 10.1 | 16.8 | 14.5 | 14 | 15 | 15 |
| sum | 6.3 | 5.9 | 7.6 | 6.9 | 9.7 | 8.7 | 9.3 | 11.4 | 11.6 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for CPTAC-5824 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
DS[+79.966331]GYYWEVPPSEVQLLK
Modification Type
Phospho (ST)
Protein - Site of Modification
N/A
Peptide - Site of Modification
3
Peptide Start
298
Peptide End
314
CPTAC ID
CPTAC-5824
Peptide Molecular Mass
2,088.9445
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
Cell Lysate
Submitting Laboratory
Fred Hutchinson Cancer Research Center, H. Lee Moffitt Cancer Center & Research Institute, Broad Institute
Submitting Lab PI
Amanda Paulovich, John Koomen, Steven A. Carr
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
SCIEX 5500 / Thermo Quantiva
Internal Standard
Stable Isotope Labeled Standard
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N K
LC
Dionex Ultimate 3000
Column Packing
C18 PepMap 100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y9 (1+) | 11.2 | 12.1 | 13.3 | 13 | 14.2 | 10.6 | 17.2 | 18.7 | 17 | 16 | 15 | 16 |
| y8 (1+) | 27.2 | 11.6 | 30.6 | 28.8 | 13.7 | 28.4 | 39.6 | 18 | 41.7 | 16 | 15 | 16 |
| sum | 12.2 | 10.5 | 19.8 | 13.3 | 13.1 | 18.1 | 18 | 16.8 | 26.8 | 16 | 15 | 16 |
Additional Resources and Comments
Assay Details for CPTAC-5823 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Sequence
DSGYYWEVPPSEVQLLK
Modification Type
unmodified
Protein - Site of Modification
N/A
Peptide - Site of Modification
3
Peptide Start
298
Peptide End
314
CPTAC ID
CPTAC-5823
Peptide Molecular Mass
2,008.9782
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
Cell Lysate
Submitting Laboratory
Fred Hutchinson Cancer Research Center, H. Lee Moffitt Cancer Center & Research Institute, Broad Institute
Submitting Lab PI
Amanda Paulovich, John Koomen, Steven A. Carr
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
SCIEX 5500 / Thermo Quantiva
Internal Standard
Stable Isotope Labeled Standard
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N K
LC
Dionex Ultimate 3000
Column Packing
C18 PepMap 100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y9 (1+) | 8.7 | 7.8 | 5.2 | 11.7 | 10.8 | 9.5 | 14.6 | 13.3 | 10.8 | 16 | 15 | 17 |
| y10 (1+) | 13.2 | 7 | 4.5 | 11.9 | 11.4 | 8.7 | 17.8 | 13.4 | 9.8 | 16 | 15 | 17 |
| sum | 9.1 | 7.6 | 4.9 | 11.5 | 10.7 | 9.4 | 14.7 | 13.1 | 10.6 | 16 | 15 | 17 |
Additional Resources and Comments
Assay Details for CPTAC-5776 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
GLNQDC[+57.021464]C[+57.021464]VVYR
Modification Type
Carbamidomethyl (C)
Protein - Site of Modification
N/A
Peptide - Site of Modification
3
Peptide Start
53
Peptide End
63
CPTAC ID
CPTAC-5776
Peptide Molecular Mass
1,382.6071
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
Cell Lysate
Submitting Laboratory
Fred Hutchinson Cancer Research Center, H. Lee Moffitt Cancer Center & Research Institute, Broad Institute
Submitting Lab PI
Amanda Paulovich, John Koomen, Steven A. Carr
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
SCIEX 5500 / Thermo Quantiva
Internal Standard
Stable Isotope Labeled Standard
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N at C-terminus R
LC
Dionex Ultimate 3000
Column Packing
C18 PepMap 100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y7 (1+) | 20 | 12.4 | 7.3 | 23.4 | 29.2 | 16.5 | 30.8 | 31.7 | 18 | 15 | 15 | 15 |
| y6 (1+) | 19.8 | 11.2 | 8.2 | 25.6 | 29.2 | 16.4 | 32.4 | 31.3 | 18.3 | 15 | 15 | 15 |
| y5 (1+) | 18.6 | 10.2 | 7.5 | 24.4 | 28 | 15.1 | 30.7 | 29.8 | 16.9 | 15 | 15 | 15 |
| sum | 16.6 | 11.4 | 7.6 | 23.4 | 28.9 | 16.1 | 28.7 | 31.1 | 17.8 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for CPTAC-5775 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Sequence
IGTGSFGTVFR
Modification Type
unmodified
Protein - Site of Modification
N/A
Peptide - Site of Modification
3
Peptide Start
316
Peptide End
326
CPTAC ID
CPTAC-5775
Peptide Molecular Mass
1,140.5928
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
Cell Lysate
Submitting Laboratory
Fred Hutchinson Cancer Research Center, H. Lee Moffitt Cancer Center & Research Institute, Broad Institute
Submitting Lab PI
Amanda Paulovich, John Koomen, Steven A. Carr
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
SCIEX 5500 / Thermo Quantiva
Internal Standard
Stable Isotope Labeled Standard
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N at C-terminus R
LC
Dionex Ultimate 3000
Column Packing
C18 PepMap 100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y5 (1+) | 18.9 | 10.5 | 6 | 22.1 | 28.2 | 12.1 | 29.1 | 30.1 | 13.5 | 15 | 15 | 15 |
| y6 (1+) | 17.7 | 8.1 | 6.7 | 21.2 | 27 | 12.5 | 27.6 | 28.2 | 14.2 | 15 | 15 | 15 |
| y8 (1+) | 20.3 | 10.3 | 6.7 | 25.3 | 27.1 | 12.6 | 32.4 | 29 | 14.3 | 15 | 15 | 15 |
| sum | 18.3 | 9.7 | 6.2 | 21.9 | 27.5 | 12.2 | 28.5 | 29.2 | 13.7 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for CPTAC-5826 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
NLGYRDS[+79.966331]GYYWEVPPSEVQLLK
Modification Type
Phospho (ST)
Protein - Site of Modification
N/A
Peptide - Site of Modification
3
Peptide Start
293
Peptide End
314
CPTAC ID
CPTAC-5826
Peptide Molecular Mass
2,692.2574
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
Cell Lysate
Submitting Laboratory
Fred Hutchinson Cancer Research Center, H. Lee Moffitt Cancer Center & Research Institute, Broad Institute
Submitting Lab PI
Amanda Paulovich, John Koomen, Steven A. Carr
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
SCIEX 5500 / Thermo Quantiva
Internal Standard
Stable Isotope Labeled Standard
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N K
LC
Dionex Ultimate 3000
Column Packing
C18 PepMap 100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y9 (1+) | 8.3 | 14.3 | 5.3 | 10.4 | 14.4 | 10.3 | 13.3 | 20.3 | 11.6 | 16 | 15 | 16 |
| y8 (1+) | 10.4 | 10.3 | 9.7 | 13.3 | 14.9 | 12.8 | 16.9 | 18.1 | 16.1 | 16 | 15 | 16 |
| y9 (2+) | 11.3 | 13.3 | 7.8 | 13.1 | 14.8 | 9.5 | 17.3 | 19.9 | 12.3 | 16 | 15 | 16 |
| sum | 9 | 13.5 | 5.9 | 11.3 | 14.6 | 9.5 | 14.4 | 19.9 | 11.2 | 16 | 15 | 16 |
Additional Resources and Comments
Assay Details for CPTAC-5825 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Sequence
NLGYRDSGYYWEVPPSEVQLLK
Modification Type
unmodified
Protein - Site of Modification
N/A
Peptide - Site of Modification
3
Peptide Start
293
Peptide End
314
CPTAC ID
CPTAC-5825
Peptide Molecular Mass
2,612.2911
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
Cell Lysate
Submitting Laboratory
Fred Hutchinson Cancer Research Center, H. Lee Moffitt Cancer Center & Research Institute, Broad Institute
Submitting Lab PI
Amanda Paulovich, John Koomen, Steven A. Carr
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
SCIEX 5500 / Thermo Quantiva
Internal Standard
Stable Isotope Labeled Standard
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N K
LC
Dionex Ultimate 3000
Column Packing
C18 PepMap 100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| b12 (2+) | 69.8 | 16 | 22.2 | 91.5 | 16.4 | 22.1 | 115.1 | 22.9 | 31.3 | 16 | 15 | 17 |
| y9 (2+) | 66.4 | 17.6 | 20.8 | 93.5 | 18.4 | 20.2 | 114.7 | 25.5 | 29 | 16 | 15 | 17 |
| y9 (1+) | 62.2 | 18.8 | 12.1 | 89.6 | 14.1 | 13.2 | 109.1 | 23.5 | 17.9 | 16 | 15 | 17 |
| sum | 66.7 | 16.8 | 13 | 90.1 | 15.2 | 13.6 | 112.1 | 22.7 | 18.8 | 16 | 15 | 17 |
Additional Resources and Comments
Assay Details for CPTAC-5774 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
TQADELPAC[+57.021464]LLSAAR
Modification Type
Carbamidomethyl (C)
Protein - Site of Modification
N/A
Peptide - Site of Modification
3
Peptide Start
589
Peptide End
603
CPTAC ID
CPTAC-5774
Peptide Molecular Mass
1,614.8036
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
Cell Lysate
Submitting Laboratory
Fred Hutchinson Cancer Research Center, H. Lee Moffitt Cancer Center & Research Institute, Broad Institute
Submitting Lab PI
Amanda Paulovich, John Koomen, Steven A. Carr
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
SCIEX 5500 / Thermo Quantiva
Internal Standard
Stable Isotope Labeled Standard
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N at C-terminus R
LC
Dionex Ultimate 3000
Column Packing
C18 PepMap 100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y10 (1+) | 39.1 | 8 | 6.7 | 42.5 | 28.7 | 13.4 | 57.7 | 29.8 | 15 | 15 | 15 | 15 |
| y9 (1+) | 18.4 | 10.3 | 6 | 30.1 | 30.4 | 14 | 35.3 | 32.1 | 15.2 | 15 | 15 | 15 |
| sum | 19.3 | 9.8 | 5.9 | 30.4 | 30.1 | 13.9 | 36 | 31.7 | 15.1 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for non-CPTAC-5730 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
DS[+79.966331]GYYWEVPPSEVQLLK
Modification Type
Phospho (ST)
Protein - Site of Modification
N/A
Peptide - Site of Modification
3
Peptide Start
298
Peptide End
314
CPTAC ID
non-CPTAC-5730
Peptide Molecular Mass
2,088.9445
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
Cell Lysate
Submitting Laboratory
Moffitt Cancer Center
Submitting Lab PI
John Koomen
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
Thermo Quantiva
Internal Standard
peptide
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N K
LC
Dionex Ultimate 3000 RSLCnano
Column Packing
Pepmap 100
Column Dimensions
75um x 25cm
Flow Rate
300 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| b7-98 (1+) | 5.9 | 5.4 | 2.5 | 8.6 | 6 | 2.8 | 10.4 | 8.1 | 3.8 | 15 | 15 | 15 |
| b6-98 (1+) | 5.1 | 5.7 | 3.3 | 6 | 5.8 | 4.1 | 7.9 | 8.1 | 5.3 | 15 | 15 | 15 |
| y9 (1+) | 2.8 | 3.3 | 3.1 | 3.6 | 4.1 | 3.8 | 4.6 | 5.3 | 4.9 | 15 | 15 | 15 |
| y10 (1+) | 4.9 | 3.5 | 3.5 | 4.7 | 3.3 | 4.4 | 6.8 | 4.8 | 5.6 | 15 | 15 | 15 |
| sum | 2.4 | 3.3 | 2.8 | 3.5 | 4.1 | 3.5 | 4.2 | 5.3 | 4.5 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for non-CPTAC-5348 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Sequence
DSGYYWEVPPSEVQLLK
Modification Type
unmodified
Protein - Site of Modification
N/A
Peptide - Site of Modification
N/A
Peptide Start
298
Peptide End
314
CPTAC ID
non-CPTAC-5348
Peptide Molecular Mass
2,008.9782
Species
Homo sapiens (Human)
Assay Type
Direct MRM
Matrix
Cell Lysate
Submitting Laboratory
Moffitt Cancer Center
Submitting Lab PI
John Koomen
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
RSLCnano and Quantiva
Internal Standard
Stable Isotope Labeled Standard
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N K
LC
RSLCnano
Column Packing
C18 PepMap 100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nl/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y8 (1+) | 33.3 | 8.7 | 9.4 | 34 | 14.4 | 11.7 | 47.6 | 16.8 | 15 | 15 | 15 | 15 |
| y9 (1+) | 4.4 | 2.4 | 2.5 | 7 | 9 | 3.7 | 8.3 | 9.3 | 4.5 | 15 | 15 | 15 |
| y10 (1+) | 17.5 | 8.5 | 5.8 | 15.1 | 9.6 | 6.8 | 23.1 | 12.8 | 8.9 | 15 | 15 | 15 |
| sum | 5.6 | 2 | 2.5 | 7.8 | 8.9 | 3.9 | 9.6 | 9.1 | 4.6 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for non-CPTAC-5346 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
GLNQDC[+57.021464]C[+57.021464]VVYR
Modification Type
Carbamidomethyl (C), Carbamidomethyl (C)
Protein - Site of Modification
59, 60
Peptide - Site of Modification
6, 7
Peptide Start
53
Peptide End
63
CPTAC ID
non-CPTAC-5346
Peptide Molecular Mass
1,382.6071
Species
Homo sapiens (Human)
Assay Type
Direct MRM
Matrix
Cell Lysate
Submitting Laboratory
Moffitt Cancer Center
Submitting Lab PI
John Koomen
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
RSLCnano and Quantiva
Internal Standard
Stable Isotope Labeled Standard
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N at C-terminus R
LC
RSLCnano
Column Packing
C18 PepMap 100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nl/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y5 (1+) | 6.4 | 3.3 | 2.2 | 6 | 3.5 | 3.7 | 8.8 | 4.8 | 4.3 | 15 | 15 | 15 |
| y6 (1+) | 5.6 | 3.5 | 1.5 | 4.8 | 3.3 | 3.1 | 7.4 | 4.8 | 3.4 | 15 | 15 | 15 |
| y7 (1+) | 7.3 | 3.4 | 1.6 | 7.7 | 4.7 | 2.8 | 10.6 | 5.8 | 3.2 | 15 | 15 | 15 |
| y8 (1+) | 7.9 | 4.6 | 2.2 | 8.1 | 4.9 | 3.8 | 11.3 | 6.7 | 4.4 | 15 | 15 | 15 |
| sum | 4.4 | 2.3 | 0.9 | 3.9 | 2.7 | 2.5 | 5.9 | 3.5 | 2.7 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for non-CPTAC-5533 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Sequence
IGTGSFGTVFR
Modification Type
unmodified
Protein - Site of Modification
N/A
Peptide - Site of Modification
6
Peptide Start
316
Peptide End
326
CPTAC ID
non-CPTAC-5533
Peptide Molecular Mass
1,140.5928
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
Cell Line
Submitting Laboratory
Moffitt Cancer Center
Submitting Lab PI
John Koomen
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
RSLCnano and Quantiva
Internal Standard
SIS Peptide
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N at C-terminus R
LC
RSLCnano
Column Packing
C18 PepMap100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y5 (1+) | 19.1 | 7.5 | 6.6 | 24.2 | 28.9 | 12.4 | 30.8 | 29.9 | 14 | 14 | 14 | 14 |
| y6 (1+) | 16.8 | 5.9 | 6.9 | 22.6 | 28.5 | 12.3 | 28.2 | 29.1 | 14.1 | 14 | 14 | 14 |
| y7 (1+) | 17.2 | 6.5 | 6.9 | 22.5 | 29.1 | 12.9 | 28.3 | 29.8 | 14.6 | 14 | 14 | 14 |
| y8 (1+) | 21.2 | 7.3 | 6.6 | 27.3 | 27.6 | 12 | 34.6 | 28.5 | 13.7 | 14 | 14 | 14 |
| sum | 16.8 | 6.8 | 6.6 | 22.3 | 28.5 | 12.3 | 27.9 | 29.3 | 14 | 14 | 14 | 14 |
Additional Resources and Comments
Assay Details for non-CPTAC-5731 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
NLGYRDS[+79.966331]GYYWEVPPSEVQLLK
Modification Type
Phospho (ST)
Protein - Site of Modification
N/A
Peptide - Site of Modification
3
Peptide Start
293
Peptide End
314
CPTAC ID
non-CPTAC-5731
Peptide Molecular Mass
2,692.2574
Species
Homo sapiens (Human)
Assay Type
Enrichment MRM
Enrichment Method
Fe(III)-NTA
Matrix
Cell Lysate
Submitting Laboratory
Moffitt Cancer Center
Submitting Lab PI
John Koomen
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
Thermo Quantiva
Internal Standard
peptide
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N K
LC
Dionex Ultimate 3000 RSLCnano
Column Packing
Pepmap 100
Column Dimensions
75um x 25cm
Flow Rate
300 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| b10 (1+) | 17.6 | 6.9 | 8.2 | 17.3 | 7.5 | 9.3 | 24.7 | 10.2 | 12.4 | 15 | 15 | 15 |
| y9 (2+) | 9 | 2.2 | 2.3 | 7 | 1.7 | 2.1 | 11.4 | 2.8 | 3.1 | 15 | 15 | 15 |
| y8 (1+) | 12.5 | 6.3 | 6.2 | 13.4 | 5.4 | 6.4 | 18.3 | 8.3 | 8.9 | 15 | 15 | 15 |
| y9 (1+) | 8.7 | 1.7 | 2.9 | 10.7 | 2.3 | 3 | 13.8 | 2.9 | 4.2 | 15 | 15 | 15 |
| sum | 7.2 | 1.5 | 1.8 | 7.3 | 1.5 | 1.8 | 10.3 | 2.1 | 2.5 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for non-CPTAC-5347 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Sequence
NLGYRDSGYYWEVPPSEVQLLK
Modification Type
unmodified
Protein - Site of Modification
N/A
Peptide - Site of Modification
N/A
Peptide Start
293
Peptide End
314
CPTAC ID
non-CPTAC-5347
Peptide Molecular Mass
2,612.2911
Species
Homo sapiens (Human)
Assay Type
Direct MRM
Matrix
Cell Lysate
Submitting Laboratory
Moffitt Cancer Center
Submitting Lab PI
John Koomen
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
RSLCnano and Quantiva
Internal Standard
Stable Isotope Labeled Standard
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N K
LC
RSLCnano
Column Packing
C18 PepMap 100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nl/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| b10 (1+) | 7.2 | 4.7 | 4.5 | 11.6 | 8.2 | 5.4 | 13.7 | 9.5 | 7 | 15 | 15 | 15 |
| b9 (1+) | 13.6 | 5.9 | 4 | 12.5 | 9.6 | 5.7 | 18.5 | 11.3 | 7 | 15 | 15 | 15 |
| y8 (1+) | 14 | 6.6 | 5.2 | 15.4 | 11.3 | 6.3 | 20.8 | 13.1 | 8.2 | 15 | 15 | 15 |
| y9 (1+) | 5.8 | 4 | 0.8 | 8 | 9.8 | 3.9 | 9.9 | 10.6 | 4 | 15 | 15 | 15 |
| sum | 5.1 | 3.8 | 0.9 | 6.8 | 9.5 | 4 | 8.5 | 10.2 | 4.1 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for non-CPTAC-5351 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
TQADELPAC[+57.021464]LLSAAR
Modification Type
Carbamidomethyl (C)
Protein - Site of Modification
598
Peptide - Site of Modification
9
Peptide Start
589
Peptide End
603
CPTAC ID
non-CPTAC-5351
Peptide Molecular Mass
1,614.8036
Species
Homo sapiens (Human)
Assay Type
Direct MRM
Matrix
Cell Lysate
Submitting Laboratory
Moffitt Cancer Center
Submitting Lab PI
John Koomen
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
RSLCnano and Quantiva
Internal Standard
Stable Isotope Labeled Standard
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N at C-terminus R
LC
RSLCnano
Column Packing
C18 PepMap 100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nl/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y9 (1+) | 3.5 | 1.2 | 1.4 | 4.2 | 2.8 | 1.5 | 5.5 | 3 | 2.1 | 15 | 15 | 15 |
| y10 (1+) | 11 | 4 | 3.7 | 13.2 | 5 | 3.9 | 17.2 | 6.4 | 5.4 | 15 | 15 | 15 |
| y12 (1+) | 13.6 | 5 | 3.4 | 16.5 | 5.1 | 5.1 | 21.4 | 7.1 | 6.1 | 15 | 15 | 15 |
| y13 (1+) | 8.3 | 3.5 | 3.9 | 6.3 | 4.6 | 4.2 | 10.4 | 5.8 | 5.7 | 15 | 15 | 15 |
| sum | 2.1 | 1 | 1.3 | 3.5 | 2.7 | 1.7 | 4.1 | 2.9 | 2.1 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for non-CPTAC-5350 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Sequence
VSQPTAEQAQAFK
Modification Type
unmodified
Protein - Site of Modification
N/A
Peptide - Site of Modification
N/A
Peptide Start
340
Peptide End
352
CPTAC ID
non-CPTAC-5350
Peptide Molecular Mass
1,403.7045
Species
Homo sapiens (Human)
Assay Type
Direct MRM
Matrix
Cell Lysate
Submitting Laboratory
Moffitt Cancer Center
Submitting Lab PI
John Koomen
Publication
View Details (opens in a new window)
Targeted mass spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP Kinase, and AKT signaling. Whiteaker JR, Sharma K, Hoffman MA, Kuhn E, Zhao L, Cocco AR, Schoenherr RM, Kennedy JJ, Voytovich U, Lin C, Fang B, Bowers K, Whiteley G, Colantonio S, Bocik W, Roberts R, Hiltke T, Boja E, Rodriguez H, McCormick F, Holderfield M, Carr SA, Koomen JM, Paulovich AG. Cell Reports Methods doi: 10.1016/j.crmeth.2021.100015.
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
RSLCnano and Quantiva
Internal Standard
Stable Isotope Labeled Standard
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N at C-terminus K
LC
RSLCnano
Column Packing
C18 PepMap 100
Column Dimensions
75 micron ID x 25 cm length
Flow Rate
300 nl/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y10 (1+) | 5.5 | 2.2 | 1.7 | 5.3 | 2.8 | 2.3 | 7.6 | 3.6 | 2.9 | 15 | 15 | 15 |
| y5 (1+) | 13.5 | 2.6 | 4.2 | 13.5 | 5.9 | 5.4 | 19.1 | 6.4 | 6.8 | 15 | 15 | 15 |
| y6 (1+) | 10.6 | 4.3 | 4.9 | 13.2 | 4.5 | 6 | 16.9 | 6.2 | 7.7 | 15 | 15 | 15 |
| y7 (1+) | 12.9 | 5.4 | 3.3 | 12.2 | 7.8 | 4.3 | 17.8 | 9.5 | 5.4 | 15 | 15 | 15 |
| y8 (1+) | 15.9 | 6.6 | 3.9 | 16.4 | 7.9 | 5.3 | 22.8 | 10.3 | 6.6 | 15 | 15 | 15 |
| sum | 4.7 | 1.8 | 1.6 | 5.3 | 3.3 | 2.3 | 7.1 | 3.8 | 2.8 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for CPTAC-1547 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Modified Sequence
TQHC[+57.0]DPEHFPFPAPANAPLQR
Modification Type
Carbamidomethyl (C)
Protein - Site of Modification
192
Peptide - Site of Modification
4
Peptide Start
189
Peptide End
209
CPTAC ID
CPTAC-1547
Peptide Molecular Mass
2,429.1335
Species
Homo sapiens (Human)
Assay Type
Direct MRM
Matrix
Ovarian cancer tumor tissue lysate
Submitting Laboratory
Pacific Northwest National Laboratory
Submitting Lab PI
Tao Liu
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
TSQ Vantage
Internal Standard
peptide
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N at C-terminus R
LC
Waters nanoACQUITY (Part Number 176016000)
Column Packing
Waters BEH C18, 1.7 um 130 Å (Part Number 186007485)
Column Dimensions
100 um x 100 mm
Flow Rate
0.5 uL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y6 (1+) | 26.1 | 3.9 | 4.8 | 26.7 | 10.3 | 12.6 | 37.3 | 11 | 13.5 | 15 | 15 | 15 |
| y4 (1+) | 15.4 | 8.1 | 5.8 | 22.8 | 10.2 | 8.9 | 27.5 | 13 | 10.6 | 15 | 15 | 15 |
| b11 (2+) | 9.7 | 3.9 | 4.1 | 25.6 | 7.1 | 7.1 | 27.4 | 8.1 | 8.2 | 15 | 15 | 15 |
| b13 (2+) | 11.4 | 3.9 | 2.8 | 20.3 | 7.8 | 7 | 23.3 | 8.7 | 7.5 | 15 | 15 | 15 |
| sum | 5.3 | 2.5 | 2.3 | 16.1 | 7.3 | 7 | 16.9 | 7.7 | 7.4 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for non-CPTAC-3958 Collapse assay details
Data source: Panorama
Official Gene Symbol
Araf
Peptide Sequence
IGDFGLATVK
Modification Type
unmodified
Protein - Site of Modification
N/A
Peptide - Site of Modification
N/A
Peptide Start
443
Peptide End
452
CPTAC ID
non-CPTAC-3958
Peptide Molecular Mass
1,019.5651
Species
Mus musculus (Mouse)
Assay Type
Direct MRM
Matrix
Plasma
Submitting Laboratory
UVic-Genome BC Proteomics Centre
Submitting Lab PI
Christoph Borchers
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
Agilent 6490/6495 QQQ
Internal Standard
synthetic peptide
Peptide Standard Purity
>80%
Peptide Standard Label Type
13C and 15N at C-terminus K
LC
1290 LC (Agilent)
Column Packing
Zorbax Eclipse Plus C18, 1.8 µm
Column Dimensions
2.1 x 150 mm
Flow Rate
400 µL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y7 (1+) | 16.1 | 3.7 | 2.5 | 16 | 5.4 | 3.4 | 22.7 | 6.5 | 4.2 | 15 | 15 | 15 |
| y8 (1+) | 5.3 | 3.2 | 2.9 | 7.9 | 4.4 | 5.5 | 9.5 | 5.4 | 6.2 | 15 | 15 | 15 |
| y9 (1+) | 8.2 | 2.4 | 2.9 | 11.9 | 4 | 5.7 | 14.5 | 4.7 | 6.4 | 15 | 15 | 15 |
| b3 (1+) | 6.1 | 2.5 | 4.4 | 7.2 | 4.9 | 5.4 | 9.4 | 5.5 | 7 | 15 | 15 | 15 |
| y6 (1+) | 7.2 | 3.4 | 2.6 | 12.5 | 4.6 | 4.3 | 14.4 | 5.7 | 5 | 15 | 15 | 15 |
| sum | 2.7 | 1.1 | 1.3 | 7.4 | 3.3 | 4 | 7.9 | 3.5 | 4.2 | 15 | 15 | 15 |
Additional Resources and Comments
Assay Details for CPTAC-3065 Collapse assay details
Data source: Panorama
Official Gene Symbol
ARAF
Peptide Sequence
VSQPTAEQAQAFK
Modification Type
unmodified
Protein - Site of Modification
N/A
Peptide - Site of Modification
N/A
Peptide Start
340
Peptide End
352
CPTAC ID
CPTAC-3065
Peptide Molecular Mass
1,403.7045
Species
Homo Sapiens
Assay Type
Direct MRM
Matrix
Tumor Digest
Submitting Laboratory
Washington University in St. Louis
Submitting Lab PI
Reid Townsend
Assay Parameters Collapse assay parameters
Data source: Panorama
Instrument
ThermoFisher, Q-Exactive
Internal Standard
25 fmol on column
Peptide Standard Purity
>95%
Peptide Standard Label Type
13C and 15N at C-terminus K
LC
Easy-nLC 1000 Thermo Scientific
Column Packing
C18
Column Dimensions
75 µm x 50 cm
Flow Rate
300 nL/min
Assay Multiplexing Expand assay panel
Broad Institute-Biologist curated set of targets across a number of important cancer pathways
Chromatograms
Data source: Panorama
Response Curves
Data source: Panorama
Retrieving Data
Repeatability
Data source: Panorama
| | Average intra-assay CV(within day CV) | Average inter-assay CV(between day CV) | Total CV | n= | | | | | | | | | |
| ---------------------------------------- | -------------------------------------- | --------------------------------------------------------------------------------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- | ------- | ------- | -------- |
| Fragment ion / Transition | Low | Med | High | Low | Med | High | Low | Med | High | Low | Med | High |
| y7 (1+) | 5.1 | 3.8 | 2.2 | 5.6 | 4.1 | 3.2 | 7.6 | 5.6 | 3.9 | 15 | 15 | 15 |
| y8 (1+) | 4.8 | 3.8 | 2.8 | 5.7 | 3.9 | 3.6 | 7.5 | 5.4 | 4.6 | 15 | 15 | 15 |
| y10 (1+) | 3.9 | 3.4 | 2.2 | 5.6 | 3.5 | 2.2 | 6.8 | 4.9 | 3.1 | 15 | 15 | 15 |
| sum | 3.6 | 3.1 | 1.9 | 5.1 | 3.3 | 2.3 | 6.2 | 4.5 | 3 | 15 | 15 | 15 |