John Hurrell - Academia.edu (original) (raw)
Papers by John Hurrell
The Medical Journal of Australia, Nov 1, 1982
in th is form. However , I believe that this form or something similar could be usefully introduc... more in th is form. However , I believe that this form or something similar could be usefully introduced. It would at least be better than the present confused situation. o. B. Wilson.
PubMed, Apr 1, 1987
A very rapid and sensitive assay for human choriogonadotropin (hCG) has been developed involving ... more A very rapid and sensitive assay for human choriogonadotropin (hCG) has been developed involving two beta-subunit-specific monoclonal antibodies. In the assay the test specimen is passed backward and forward (reflow) through a monoclonal-antibody-coated capillary tube for 1 min, then incubated for 1 min with a second monoclonal antibody conjugated to urease (EC 3.5.1.5). After addition of a urease substrate solution, 10 int. units of hCG per liter can be detected visually within 5 min, which compares very favorably with other currently available hCG assay procedures. Advantages of the reflow/capillary tube assay system and optimization of the test procedure are discussed.
PubMed, Sep 1, 1981
Hybridoma cell lines secreting monoclonal antibodies to carcinoembryonic antigen (CEA) were gener... more Hybridoma cell lines secreting monoclonal antibodies to carcinoembryonic antigen (CEA) were generated by fusing mouse immune lymphocytes with the mouse myeloma variant cell line, NS-1. Antibody secreted by one cloned hybrid cell line could bind only a select portion of the CEA bound by the commercially available goat anti-CEA antiserum used in clinical assays. Radiolabeled CEA could be purified on a monoclonal antibody affinity column. Incorporation of this purified radiolabeled CEA in a double-antibody solid-phase assay with goat anti-CEA antiserum led to an approximately 2.5-fold increase in sensitivity of the assay. Genetically stable hybrid clones may be sources of virtually unlimited quantities of such antibodies which may have potential utility in improving the cancer specificity of clinical assays.
Federation Proceedings, Jul 29, 1980
Journal of Biological Chemistry, Mar 1, 1982
Elsevier eBooks, 1994
This chapter presents biosensors in diagnostics. The healthcare industry in the United States has... more This chapter presents biosensors in diagnostics. The healthcare industry in the United States has been undergoing significant change over the past three years that has provided both, opportunity and barriers to the application of biosensors in diagnostics. Productivity and cost containment pressures have made diagnostic laboratories particularly sensitive to the value of new technology. Capitation contracts to managed care organizations have made many laboratories walk a very thin line between profit and loss. The advent of broader testing and screening for genetic based disorders provides another opportunity for biosensor technology. The discovery of the multiple mutations involved with many disorders, such as cystic fibrosis and the multigenic basis of cancer and other diseases, makes conventional testing difficult and expensive. Biosensor technology may provide the answer to rapid and inexpensive genetic testing in the future. Biosensors have some unique attributes that make them particularly suited for use in sites requiring fast test results and limited sample preparation.
Current protocols in immunology, Jun 1, 1998
Immunoblotting (often referred to as western blotting) is used to identify specific antigens reco... more Immunoblotting (often referred to as western blotting) is used to identify specific antigens recognized by polyclonal or monoclonal antibodies. This unit presents procedures for electrophoretically transferring antigens from a denaturing polyacrylamide gel in a tank or a semidry transfer apparatus to a nitrocellulose, PVDF, or nylon membrane. The process can be monitored by reversible staining or by Ponceau S staining, both of which are described here. A protocol for blotting previously stained gels is also described. The transferred proteins are bound to the surface of the membrane, providing access for reaction with immunodetection reagents. All remaining binding sites are blocked by immersing the membrane in a solution containing either a protein or detergent blocking agent. After probing with the primary antibody, the membrane is washed and the antibody‐antigen complexes are identified with horseradish peroxidase (HRPO) or alkaline phosphatase enzymes coupled to the secondary anti‐IgG antibody (e.g., goat anti‐rabbit IgG). The enzymes are attached directly or via an avidin‐biotin bridge to the secondary antibody, and protocols are provided for both methods. Chromogenic or luminescent substrates are then used as described to visualize the activity. Finally, a method for stripping and reprobing membranes is presented.
Molecular Immunology, Mar 1, 1982
In some instances. even the increased resolutton that may be afforded in immunoassays by the use ... more In some instances. even the increased resolutton that may be afforded in immunoassays by the use of monoclonal antibodies fails to effect resolution among molecules that share many epitopes. An immunoradiometric assay that simultaneously measured two different epitopes on the same molecule was devised to overcome this difhculty in the differentiation between cardiac-and skeletal-myosin light chains. Three monoclonal antibodies were examined that were loo", (lC5). 25"" (2B9) and 17"" (4FlO) cross reactive. respectively. between the two antigens. One antibody of the pair to be studied was immobilized to cyanogen bromide-activated Sepharose 4B while the other was iodinated with "'1 using the lactoperoxidase method. The antigen was mixed with the immobilized antibody. the labeled antibody was added and the precipitate then washed and counted in a gamma counter. When both antibodies of the pair to be studied (immobtlized and labeled) were the same (2B9). no radioactivity above background was bound to the precipitate. indicating that the second antibody could not bind to an already occupied epitope. When two different antibodies were employed. the specificity of the assay increased over that of a single antibody. The cross reactivity of a pair approximated the product of the cross reacttvities of the individual antibodies. Thus. lC5 and 2B9 were X,, cross reactive together. lC5 and 4FlO 17",, cross reactive. and 3B9 and 4FIO 4.3",, cross reactive.
Journal of Immunological Methods, Sep 1, 1981
Spleen cells from BALB/c mice, immunized with SDS-acrylamide gel purified human fibroblast interf... more Spleen cells from BALB/c mice, immunized with SDS-acrylamide gel purified human fibroblast interferon (HuIFN-ß) were fused with mouse myeloma cells. Culture fluids from the resulting hybrid cells were screened for HuIFN-ß specificity by the following tests: (1) a solid-phase RIA using partially purified HuIFN-0, (2) a protein-transfer RIA using electrophoretically resolved and immobilized HuIF\-/3, and (3) a bioassay which tests residual HuIFN-/3 activity in the supernatant following immunoprecipitation. Six HuIFN-/3-specific hybridomas were identified; two were capable of partially neutralizing HuIFN-j3 activity. All six antibodies bind to the /31-IFN polypeptide synthesized in E. coli cells containing a cloned /31-IFN DNA sequence. All six monoclonal antibodies were found to be IgG3/x.
Biochemistry, Oct 1, 1975
An improved separation procedure is described for isolating five leghemoglobin components from th... more An improved separation procedure is described for isolating five leghemoglobin components from the nodules of soybean plants. After a preliminary oxidation with ferricyanide, and separation from endogenous nicotinate at pH 9.2, the ferrileghemoglobins are separated by DEAE-cellulose chromatography using gradient elution with acetate buffer (pH 5.2). The components have been characterized by their acetate and nicotinate binding affinities, gel electrophoretic, visible, and circular dichroic spectra in the ultraviolet, Soret and visible regions. Two formerly unresolved components of leghemoglobin c have indistinguishable circular dichroic, electrophoretic, and ligand binding properties, but differ in their spin states as judged by their visible spectra, their amino acid analyses, and their tryptic maps.
Journal of Infection, Mar 1, 1984
A sensitive enzyme immunoassay for rapidly assessing a patient's state of immunity to tetanus is ... more A sensitive enzyme immunoassay for rapidly assessing a patient's state of immunity to tetanus is described. The test, which uses 50 ~tl sample of blood, plasma or serum, is done in a capillary tube and, by comparison with two adjacent reference tubes containing standardised sera, places immunity to tetanus in one of three categorieslow-negative (<o-oi IU/ml), intermediate (o-oi-I.28 IU/ml) or high (> 1.28 IU/ml). In a study of 90 clinical specimens assayed both by toxin neutralisation bioassay and capillary enzyme immunoassay the enzyme immunoassay accurately assessed the state of immunity to tetanus of the patients concerned.
Journal of Biological Standardization, 1983
Antibodies ¢o tetanus toxin were induced in sheep by hyperimmunizar~on over 24 g'ecks. BIc-eds ta... more Antibodies ¢o tetanus toxin were induced in sheep by hyperimmunizar~on over 24 g'ecks. BIc-eds taken at weeks 4.8.20 and tO were assayed for antibody titre by tx)th an enzyme immt, noass.iy (EIA) using a newly-described urease enzyme/substrate system and by bio.Lssay in mice. There g'as a very good correlation between the two assay systems and. x~-it h the exccptiLm of tht-wct-k-1 bleeds, the relationship was the same at all stages of hHx'rimmunization re/~.irdlcss of tttre. adjuvant, or whether toxin or toxoid was used as immunogen or for coatin.t: the plates. The results c-stablish that the EIA can replace the bioassay for the determination of tetanus ~lnt itoxin in ovine sera.
Journal of Immunological Methods, Sep 1, 1982
The development of urease (E.C.3.5.1.5) as a label for enzyme immunoassay (EIA) procedures is des... more The development of urease (E.C.3.5.1.5) as a label for enzyme immunoassay (EIA) procedures is described and the use of such conjugates illustrated with examples. Urease catalyzes the hydrolysis of urea to carbon dioxide and ammonia. The production of ammonia may be detected readily by a pH shift which we have found best indicated by the vivid colour change (yellow to purple) of bromocresol purple incorporated in the substrate solution. This enzyme-substrate system offers a number of important advantages. The substrate in aqueous solution is stable, titration end points are sharp and readily visible and the enzyme is not inhibited by sodium azide. Thus, test reagents may be prepared with this preservative and stored ready to use. Urease of high specific activity is commercially available and because it does not occur in mammalian tissues, it is suitable for use in EIA tests to detect cell-associated antigens and their antibodies. Finally, the enzyme reaction may be stopped by the addition of organomercurial preservatives, thus allowing storage of developed tests for later examination.
Current protocols in molecular biology, 1988
Hybridomas to be cloned are diluted to 0.8 cells/well. This dilution provides 36% of wells with 1... more Hybridomas to be cloned are diluted to 0.8 cells/well. This dilution provides 36% of wells with 1 cell/well by Poisson statistics. When cultures are 10 to 50% confluent, antibody is assayed by ELISA. Two or more cloning procedures are carried out until >90% of the wells containing single clones are positive for antibody production. Materials HAT medium (UNIT 11.6) HT medium (UNIT 11.7) Complete culture medium (UNIT 11.5) Polyvinyl or polystyrene 96-well microtiter plates 6-and 24-well culture plates 8% CO 2-in-air gas mixture Humidified CO 2 incubator Cryotubes (Nunc #3-63401) 2-, 5-, and 10-ml serological pipets Multichannel pipet and tips Inverted microscope Additional reagents and equipment for preparing mouse feeder cells for fusion and cloning (UNIT 11.6), for ELISA screening (UNIT 11.4), and for estimating cell viability by trypan blue exclusion (UNIT 11.5) 3. Perform cell viability count using trypan blue exclusion method (support protocol, UNIT 11.5) on the overnight cultures in 24-well plates. 4. Using a 6-well plate, make dilutions of cells from overnight cultures in HT or complete medium. In the first well, make a 1:100 dilution in a total of 3 ml; in the second well, dilute an aliquot of the first dilution to 80 cells/ml in 5 ml; in the third well, prepare 8 cells/ml in 10 ml (i.e., a 1:10 dilution from the second well). Choice of medium is discussed in step 1. Using 6-well plates is far easier for preparing cell dilutions than using tubes. However, using 6-well plates is expensive. 5. With a multichannel pipet, fill the upper 50 wells of the inner 60 wells of the 96-well plate from step 1 with 100 µl of 8 cells/ml dilution (i.e., 0.8 cells/well)
Biochemistry, Jan 25, 1977
The conformational motilities of three regions of the sperm whale myoglobin molecule and of an is... more The conformational motilities of three regions of the sperm whale myoglobin molecule and of an isolated peptide of myoglobin have been examined by measuring the equilibrium constant for the nativenonnative transition. The immunological approach of Furie et al.
Journal of Clinical Microbiology, Nov 1, 1983
Journal of Immunology, 1977
The use of Freund's complete adjuvant for immunizing goats with myoglobin produces mainly... more The use of Freund's complete adjuvant for immunizing goats with myoglobin produces mainly antibodies directed against antigenic determinants present in the native protein. Only about 9% of the total antibodies produced are directed toward determinants not expressed in tha native molecule. This shows that neither emulsification nor the subsequent in vivo events leading up to the immune response appreciably perturb the conformation of the protein surface.
Analytical Biochemistry, Jul 1, 1978
ABSTRACT Nonspecific adsorption of serum proteins occurs with immunoadsorption of antibodies on S... more ABSTRACT Nonspecific adsorption of serum proteins occurs with immunoadsorption of antibodies on Sepharose-myoglobin and Sepharose-staphylococcal nuclease immunoadsorbents. This adsorption results from nonspecific hydrophobic and ionic interactions between these serum proteins and the immunoadsorbents. Various preelution washing procedures were examined, and only borate-saline buffer (pH 8,5) containing a nonionic detergent, Tween 20 (0,1%), and a high salt concentration (1 m NaCl) eliminated or significantly reduced nonspecific adsorption without appreciably diminishing the recovery of specifically adsorbed antibodies.
The Medical Journal of Australia, Nov 1, 1982
in th is form. However , I believe that this form or something similar could be usefully introduc... more in th is form. However , I believe that this form or something similar could be usefully introduced. It would at least be better than the present confused situation. o. B. Wilson.
PubMed, Apr 1, 1987
A very rapid and sensitive assay for human choriogonadotropin (hCG) has been developed involving ... more A very rapid and sensitive assay for human choriogonadotropin (hCG) has been developed involving two beta-subunit-specific monoclonal antibodies. In the assay the test specimen is passed backward and forward (reflow) through a monoclonal-antibody-coated capillary tube for 1 min, then incubated for 1 min with a second monoclonal antibody conjugated to urease (EC 3.5.1.5). After addition of a urease substrate solution, 10 int. units of hCG per liter can be detected visually within 5 min, which compares very favorably with other currently available hCG assay procedures. Advantages of the reflow/capillary tube assay system and optimization of the test procedure are discussed.
PubMed, Sep 1, 1981
Hybridoma cell lines secreting monoclonal antibodies to carcinoembryonic antigen (CEA) were gener... more Hybridoma cell lines secreting monoclonal antibodies to carcinoembryonic antigen (CEA) were generated by fusing mouse immune lymphocytes with the mouse myeloma variant cell line, NS-1. Antibody secreted by one cloned hybrid cell line could bind only a select portion of the CEA bound by the commercially available goat anti-CEA antiserum used in clinical assays. Radiolabeled CEA could be purified on a monoclonal antibody affinity column. Incorporation of this purified radiolabeled CEA in a double-antibody solid-phase assay with goat anti-CEA antiserum led to an approximately 2.5-fold increase in sensitivity of the assay. Genetically stable hybrid clones may be sources of virtually unlimited quantities of such antibodies which may have potential utility in improving the cancer specificity of clinical assays.
Federation Proceedings, Jul 29, 1980
Journal of Biological Chemistry, Mar 1, 1982
Elsevier eBooks, 1994
This chapter presents biosensors in diagnostics. The healthcare industry in the United States has... more This chapter presents biosensors in diagnostics. The healthcare industry in the United States has been undergoing significant change over the past three years that has provided both, opportunity and barriers to the application of biosensors in diagnostics. Productivity and cost containment pressures have made diagnostic laboratories particularly sensitive to the value of new technology. Capitation contracts to managed care organizations have made many laboratories walk a very thin line between profit and loss. The advent of broader testing and screening for genetic based disorders provides another opportunity for biosensor technology. The discovery of the multiple mutations involved with many disorders, such as cystic fibrosis and the multigenic basis of cancer and other diseases, makes conventional testing difficult and expensive. Biosensor technology may provide the answer to rapid and inexpensive genetic testing in the future. Biosensors have some unique attributes that make them particularly suited for use in sites requiring fast test results and limited sample preparation.
Current protocols in immunology, Jun 1, 1998
Immunoblotting (often referred to as western blotting) is used to identify specific antigens reco... more Immunoblotting (often referred to as western blotting) is used to identify specific antigens recognized by polyclonal or monoclonal antibodies. This unit presents procedures for electrophoretically transferring antigens from a denaturing polyacrylamide gel in a tank or a semidry transfer apparatus to a nitrocellulose, PVDF, or nylon membrane. The process can be monitored by reversible staining or by Ponceau S staining, both of which are described here. A protocol for blotting previously stained gels is also described. The transferred proteins are bound to the surface of the membrane, providing access for reaction with immunodetection reagents. All remaining binding sites are blocked by immersing the membrane in a solution containing either a protein or detergent blocking agent. After probing with the primary antibody, the membrane is washed and the antibody‐antigen complexes are identified with horseradish peroxidase (HRPO) or alkaline phosphatase enzymes coupled to the secondary anti‐IgG antibody (e.g., goat anti‐rabbit IgG). The enzymes are attached directly or via an avidin‐biotin bridge to the secondary antibody, and protocols are provided for both methods. Chromogenic or luminescent substrates are then used as described to visualize the activity. Finally, a method for stripping and reprobing membranes is presented.
Molecular Immunology, Mar 1, 1982
In some instances. even the increased resolutton that may be afforded in immunoassays by the use ... more In some instances. even the increased resolutton that may be afforded in immunoassays by the use of monoclonal antibodies fails to effect resolution among molecules that share many epitopes. An immunoradiometric assay that simultaneously measured two different epitopes on the same molecule was devised to overcome this difhculty in the differentiation between cardiac-and skeletal-myosin light chains. Three monoclonal antibodies were examined that were loo", (lC5). 25"" (2B9) and 17"" (4FlO) cross reactive. respectively. between the two antigens. One antibody of the pair to be studied was immobilized to cyanogen bromide-activated Sepharose 4B while the other was iodinated with "'1 using the lactoperoxidase method. The antigen was mixed with the immobilized antibody. the labeled antibody was added and the precipitate then washed and counted in a gamma counter. When both antibodies of the pair to be studied (immobtlized and labeled) were the same (2B9). no radioactivity above background was bound to the precipitate. indicating that the second antibody could not bind to an already occupied epitope. When two different antibodies were employed. the specificity of the assay increased over that of a single antibody. The cross reactivity of a pair approximated the product of the cross reacttvities of the individual antibodies. Thus. lC5 and 2B9 were X,, cross reactive together. lC5 and 4FlO 17",, cross reactive. and 3B9 and 4FIO 4.3",, cross reactive.
Journal of Immunological Methods, Sep 1, 1981
Spleen cells from BALB/c mice, immunized with SDS-acrylamide gel purified human fibroblast interf... more Spleen cells from BALB/c mice, immunized with SDS-acrylamide gel purified human fibroblast interferon (HuIFN-ß) were fused with mouse myeloma cells. Culture fluids from the resulting hybrid cells were screened for HuIFN-ß specificity by the following tests: (1) a solid-phase RIA using partially purified HuIFN-0, (2) a protein-transfer RIA using electrophoretically resolved and immobilized HuIF\-/3, and (3) a bioassay which tests residual HuIFN-/3 activity in the supernatant following immunoprecipitation. Six HuIFN-/3-specific hybridomas were identified; two were capable of partially neutralizing HuIFN-j3 activity. All six antibodies bind to the /31-IFN polypeptide synthesized in E. coli cells containing a cloned /31-IFN DNA sequence. All six monoclonal antibodies were found to be IgG3/x.
Biochemistry, Oct 1, 1975
An improved separation procedure is described for isolating five leghemoglobin components from th... more An improved separation procedure is described for isolating five leghemoglobin components from the nodules of soybean plants. After a preliminary oxidation with ferricyanide, and separation from endogenous nicotinate at pH 9.2, the ferrileghemoglobins are separated by DEAE-cellulose chromatography using gradient elution with acetate buffer (pH 5.2). The components have been characterized by their acetate and nicotinate binding affinities, gel electrophoretic, visible, and circular dichroic spectra in the ultraviolet, Soret and visible regions. Two formerly unresolved components of leghemoglobin c have indistinguishable circular dichroic, electrophoretic, and ligand binding properties, but differ in their spin states as judged by their visible spectra, their amino acid analyses, and their tryptic maps.
Journal of Infection, Mar 1, 1984
A sensitive enzyme immunoassay for rapidly assessing a patient's state of immunity to tetanus is ... more A sensitive enzyme immunoassay for rapidly assessing a patient's state of immunity to tetanus is described. The test, which uses 50 ~tl sample of blood, plasma or serum, is done in a capillary tube and, by comparison with two adjacent reference tubes containing standardised sera, places immunity to tetanus in one of three categorieslow-negative (<o-oi IU/ml), intermediate (o-oi-I.28 IU/ml) or high (> 1.28 IU/ml). In a study of 90 clinical specimens assayed both by toxin neutralisation bioassay and capillary enzyme immunoassay the enzyme immunoassay accurately assessed the state of immunity to tetanus of the patients concerned.
Journal of Biological Standardization, 1983
Antibodies ¢o tetanus toxin were induced in sheep by hyperimmunizar~on over 24 g'ecks. BIc-eds ta... more Antibodies ¢o tetanus toxin were induced in sheep by hyperimmunizar~on over 24 g'ecks. BIc-eds taken at weeks 4.8.20 and tO were assayed for antibody titre by tx)th an enzyme immt, noass.iy (EIA) using a newly-described urease enzyme/substrate system and by bio.Lssay in mice. There g'as a very good correlation between the two assay systems and. x~-it h the exccptiLm of tht-wct-k-1 bleeds, the relationship was the same at all stages of hHx'rimmunization re/~.irdlcss of tttre. adjuvant, or whether toxin or toxoid was used as immunogen or for coatin.t: the plates. The results c-stablish that the EIA can replace the bioassay for the determination of tetanus ~lnt itoxin in ovine sera.
Journal of Immunological Methods, Sep 1, 1982
The development of urease (E.C.3.5.1.5) as a label for enzyme immunoassay (EIA) procedures is des... more The development of urease (E.C.3.5.1.5) as a label for enzyme immunoassay (EIA) procedures is described and the use of such conjugates illustrated with examples. Urease catalyzes the hydrolysis of urea to carbon dioxide and ammonia. The production of ammonia may be detected readily by a pH shift which we have found best indicated by the vivid colour change (yellow to purple) of bromocresol purple incorporated in the substrate solution. This enzyme-substrate system offers a number of important advantages. The substrate in aqueous solution is stable, titration end points are sharp and readily visible and the enzyme is not inhibited by sodium azide. Thus, test reagents may be prepared with this preservative and stored ready to use. Urease of high specific activity is commercially available and because it does not occur in mammalian tissues, it is suitable for use in EIA tests to detect cell-associated antigens and their antibodies. Finally, the enzyme reaction may be stopped by the addition of organomercurial preservatives, thus allowing storage of developed tests for later examination.
Current protocols in molecular biology, 1988
Hybridomas to be cloned are diluted to 0.8 cells/well. This dilution provides 36% of wells with 1... more Hybridomas to be cloned are diluted to 0.8 cells/well. This dilution provides 36% of wells with 1 cell/well by Poisson statistics. When cultures are 10 to 50% confluent, antibody is assayed by ELISA. Two or more cloning procedures are carried out until >90% of the wells containing single clones are positive for antibody production. Materials HAT medium (UNIT 11.6) HT medium (UNIT 11.7) Complete culture medium (UNIT 11.5) Polyvinyl or polystyrene 96-well microtiter plates 6-and 24-well culture plates 8% CO 2-in-air gas mixture Humidified CO 2 incubator Cryotubes (Nunc #3-63401) 2-, 5-, and 10-ml serological pipets Multichannel pipet and tips Inverted microscope Additional reagents and equipment for preparing mouse feeder cells for fusion and cloning (UNIT 11.6), for ELISA screening (UNIT 11.4), and for estimating cell viability by trypan blue exclusion (UNIT 11.5) 3. Perform cell viability count using trypan blue exclusion method (support protocol, UNIT 11.5) on the overnight cultures in 24-well plates. 4. Using a 6-well plate, make dilutions of cells from overnight cultures in HT or complete medium. In the first well, make a 1:100 dilution in a total of 3 ml; in the second well, dilute an aliquot of the first dilution to 80 cells/ml in 5 ml; in the third well, prepare 8 cells/ml in 10 ml (i.e., a 1:10 dilution from the second well). Choice of medium is discussed in step 1. Using 6-well plates is far easier for preparing cell dilutions than using tubes. However, using 6-well plates is expensive. 5. With a multichannel pipet, fill the upper 50 wells of the inner 60 wells of the 96-well plate from step 1 with 100 µl of 8 cells/ml dilution (i.e., 0.8 cells/well)
Biochemistry, Jan 25, 1977
The conformational motilities of three regions of the sperm whale myoglobin molecule and of an is... more The conformational motilities of three regions of the sperm whale myoglobin molecule and of an isolated peptide of myoglobin have been examined by measuring the equilibrium constant for the nativenonnative transition. The immunological approach of Furie et al.
Journal of Clinical Microbiology, Nov 1, 1983
Journal of Immunology, 1977
The use of Freund's complete adjuvant for immunizing goats with myoglobin produces mainly... more The use of Freund's complete adjuvant for immunizing goats with myoglobin produces mainly antibodies directed against antigenic determinants present in the native protein. Only about 9% of the total antibodies produced are directed toward determinants not expressed in tha native molecule. This shows that neither emulsification nor the subsequent in vivo events leading up to the immune response appreciably perturb the conformation of the protein surface.
Analytical Biochemistry, Jul 1, 1978
ABSTRACT Nonspecific adsorption of serum proteins occurs with immunoadsorption of antibodies on S... more ABSTRACT Nonspecific adsorption of serum proteins occurs with immunoadsorption of antibodies on Sepharose-myoglobin and Sepharose-staphylococcal nuclease immunoadsorbents. This adsorption results from nonspecific hydrophobic and ionic interactions between these serum proteins and the immunoadsorbents. Various preelution washing procedures were examined, and only borate-saline buffer (pH 8,5) containing a nonionic detergent, Tween 20 (0,1%), and a high salt concentration (1 m NaCl) eliminated or significantly reduced nonspecific adsorption without appreciably diminishing the recovery of specifically adsorbed antibodies.