Steven Shire - Academia.edu (original) (raw)

Papers by Steven Shire

Biochemistry, 1992

An expression plasmid encoding the extracellular portion of the human tumor necrosis factor (TNF)... more An expression plasmid encoding the extracellular portion of the human tumor necrosis factor (TNF) type 1 receptor (TNF-R1) was constructed and used to generate a stable cell line secreting soluble TNF-R1 (sTNF-R1). The sTNF-R1 was purified, and its biochemical properties and its interactions with human TNF-alpha were examined. SDS-PAGE resolved the purified sTNF-R1 into three bands of approximate Mr 24,200, 28,200, and 32,800. Sedimentation equilibrium analysis gave a molecular weight of 25,000 for sTNF-R1 whereas the molecular weight obtained by gel filtration chromatography was approximately 55,000-60,000. Scatchard analysis of [125I]TNF-alpha binding to sTNF-R1 revealed high-affinity binding (Kd = 93 pM), comparable to that observed for the intact receptor on whole cells. Competitive binding experiments showed that sTNF-R1 has a 50-60-fold higher affinity for TNF-alpha than for TNF-beta, in contrast to the equal affinities of TNF-alpha and TNF-beta for the full-length TNF-R1 transiently expressed in mammalian cells. The sTNF-R1 was found to block the cytotoxicity of TNF-alpha and TNF-beta on a murine L-M cell assay. The sizes of the sTNF-R1.TNF-alpha complex determined by gel filtration chromatography and sedimentation equilibrium were approximately 141 and 115 kDa, respectively. The stoichiometry of the complex was examined by Scatchard analysis, size-exclusion chromatography, HPLC separation, amino acid composition, sequence analysis, and sedimentation equilibrium. The data from these studies suggest that at least two molecules of sTNF-R1 can bind to a single TNF-alpha trimer.(ABSTRACT TRUNCATED AT 250 WORDS)

CRC critical reviews in biochemistry

Recent improvements in the understanding of electrostatic interactions in proteins serve as a foc... more Recent improvements in the understanding of electrostatic interactions in proteins serve as a focus for the general topic of pH-dependent processes in proteins. The general importance of pH-dependent processes is first set out in terms of hydrogen ion equilibria, stability, ligand interactions, assembly, dynamics, and events in related molecular systems. The development of various theoretical treatments includes various formalisms in addition to the solvent interface model developed by Shire et al. as an extension of the Tanford-Kirkwood treatment. A number of detailed applications of the model are presented and future potentialities are sketched.

... Auteur(s) / Author(s). CIPOLLA DC ; CLARK AR ; CHAN H.-K. ; GONDA I. ; SHIRE SJ ; Affiliation... more ... Auteur(s) / Author(s). CIPOLLA DC ; CLARK AR ; CHAN H.-K. ; GONDA I. ; SHIRE SJ ; Affiliation(s) du ou des auteurs / Author(s) Affiliation(s). Genentech Inc., South San Francisco CA 94080, ETATS-UNIS ... Langue / Language. Anglais Revue : Français Editeur / Publisher. ...

Journal of Biological Chemistry

A filter binding assay was developed to study interactions between purified TRAP, the trp RNA-bin... more A filter binding assay was developed to study interactions between purified TRAP, the trp RNA-binding attenuation protein of Bacillus subtilis, and trp specific transcripts. TRAP formed stable complexes with trpEDCFBA leader RNA; binding was L-tryptophan-dependent and was complete within 60 s. TRAP binds to a segment of the trp leader transcript that includes part of an RNA antiterminator structure. Binding to this segment allows formation of an RNA terminator structure, thereby promoting transcription termination. Using several trpEDCFBA leader deletion transcripts, we identified several closely spaced trinucleotide repeats (seven GAG and four UAG repeats) in the trp leader transcript that appeared to be required for TRAP binding. We also showed that TRAP binds to a segment of the trpG transcript that includes the trpG ribosome binding site; the nucleotide sequence of this segment contains several appropriately spaced trinucleotide repeats (seven GAG, one UAG, and one AAG). TRAP binding to the trpG transcript would block translation initiation. RNA footprint analysis confirmed interaction between TRAP and the trinucleotide repeats in the various transcripts. TRAP, in the presence or absence of L-tryptophan, appears to consist of 11 or 12 identical 8-kDa subunits. Our findings suggest that each tryptophan-activated TRAP subunit can bind one G/UAG repeat in a target transcript. Multiple protein-RNA interactions are required for stable association.

Ultrasensitive Biochemical Diagnostics II, 1997

ABSTRACT The use of analytical ultracentrifugation for probing macromolecular interactions in rec... more ABSTRACT The use of analytical ultracentrifugation for probing macromolecular interactions in recombinant deoxyribonucleic acid (DNA) technology was discussed. Analytical ultracentrifugation was used to determine absolute molecular weights without the use of molecular weight standards or interference from the sieving matrix used for separation. The instrument used for analytical ultracentrifugation comprised of an ultracentrifuge, designed to centrifuge components in a well controlled environment with good temperature control and a stable centrifugal field.

Monoclonal antibodies (MAb) have become a crucial therapeutic agent. However, some applications s... more Monoclonal antibodies (MAb) have become a crucial therapeutic agent. However, some applications such as a subcutaneous injection need highly concentrated protein solutions that can have undesired large viscosity. Therefore, it is of great importance to examine the relation between MAb self-associated nanostructure and its viscosity under various concentration and excipient conditions with techniques which can probe both structure and dynamics that span the length scale of an individual protein to a larger aggregate. We have investigated the protein-protein interactions and diffusive properties of highly concentrated MAbs using small angle neutron scattering (SANS) and neutron spin echo (NSE). Our results indicate that concentration, temperature, pH and surfactant do not have a strong effect on the individual MAb conformation for our MAb samples. The short-range attraction of some MAb proteins is found to be highly anisotropic in contrast to many other protein solutions at low ionic ...

The journal of physical chemistry. B, Jan 14, 2010

Light scattering intensity measurements of solutions of two purified monoclonal antibodies were p... more Light scattering intensity measurements of solutions of two purified monoclonal antibodies were performed over a wide range of concentrations (0.5-275 mg/mL) and ionic strengths (0.02 to 0.6 M). Despite extensive sequence homology between these mAbs, alteration of ∼20 amino acids in the complementarity determining regions resulted in different net intermolecular interactions and responses to solution ionic strength. The concentration dependence of scattering was analyzed by comparison with the predictions of three models, allowing for intermolecular interaction of various types. In order of increasing complexity, the three models account for: (1) steric repulsions (simple hard-sphere model), (2) steric repulsion with short-ranged attractive interactions of varying magnitude (adhesive hard-sphere model), and (3) steric and nonsteric repulsive interactions between several species whose relative concentrations may change as a function of total protein concentration as dictated by equil...

Pharmaceutical research, 2001

To determine the effect of moisture and the role of the glass transition temperature (Tg) on the ... more To determine the effect of moisture and the role of the glass transition temperature (Tg) on the stability of a high concentration, lyophilized, monoclonal antibody. A humanized monoclonal antibody was lyophilized in a sucrose/histidine/polysorbate 20 formulation. Residual moistures were from 1 to 8%. Tg values were measured by modulated DSC. Vials were stored at temperatures from 5 to 50 degrees C for 6 or 12 months. Aggregation was monitored by size exclusion chromatography and Asp isomerization by hydrophobic interaction chromatography. Changes in secondary structure were monitored by Fourier transform infrared (FTIR). T. values varied from 80 degrees C at 1% moisture to 25 degrees C at 8% moisture, there was no cake collapse and were no differences in the secondary structure by FTIR. All formulations were stable at 5 degrees C. High moisture cakes had higher aggregation rates than drier samples if stored above their Tg values. Intermediate moisture vials were more stable to aggr...

Pharmaceutical research, 1999

The purpose of this work is to utilize electron paramagnetic resonance (EPR) spectroscopy in conj... more The purpose of this work is to utilize electron paramagnetic resonance (EPR) spectroscopy in conjunction with analytical ultracentrifugation (AUC) to investigate the binding of surfactants to proteins with a transmembrance domain. As an example these methods have been used to study the interaction of a nonionic surfactant, C12E8, to recombinant human tissue factor (rhTF) in liquid formulations. The complementary nature of the two techniques aids in data interpretation when there is ambiguity using a single technique. In addition to binding stoichiometries, the possibility of identifying the interacting domains by using two forms of rhTF is explored. Two recombinant, truncated forms of human tissue factor were formulated in the absence of phospholipids. Neither of the recombinant proteins, produced in E. coli, contains the cytoplasmic domain. Recombinant human tissue factor 243 (rhTF 243) consists of 243 amino acids and includes the transmembrane sequences. Recombinant human tissue f...

Pharmaceutical research, 1999

To study the effect of trehalose, lactose, and mannitol on the biochemical stability and aerosol ... more To study the effect of trehalose, lactose, and mannitol on the biochemical stability and aerosol performance of spray-dried powders of an anti-IgE humanized monoclonal antibody. Protein aggregation of spray-dried powders stored at various temperature and relative humidity conditions was assayed by size exclusion chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis. Protein glycation was determined by isoelectric focusing and affinity chromatography. Crystallization was examined by X-ray powder diffraction. Aerosol performance was assessed as the fine particle fraction (FPF) of the powders blended with coarse carrier lactose, and was determined using a multiple stage liquid impinger. Soluble protein aggregation consisting of non-covalent and disulfide-linked covalent dimers and trimers occurred during storage. Aggregate was minimized by formulation with trehalose at or above a molar ratio in the range of 300: 1 to 500:1 (excipient:protein). However, the powder...

Pharmaceutical research, 1999

To develop a new technique, spray freeze drying, for preparing protein aerosol powders. Also, to ... more To develop a new technique, spray freeze drying, for preparing protein aerosol powders. Also, to compare the spray freeze-dried powders with spray-dried powders in terms of physical properties and aerosol performance. Protein powders were characterized using particle size analysis, thermogravimetric analysis, scanning electron microscopy, X-ray powder diffractometry, and specific surface area measurement. Aerosol performance of the powders was evaluated after blending with lactose carriers using a multi-stage liquid impinger or an Anderson cascade impactor. Two recombinant therapeutic proteins currently used for treating respiratory tract-related diseases, deoxyribonuclase (rhDNase) and anti-IgE monoclonal antibody (anti-IgE MAb), were employed and formulated with different carbohydrate excipients. Through the same atomization but the different drying process, spray drying (SD) produced small (approximately 3 microns), dense particles, but SFD resulted in large (approximately 8-10 m...

Pharmaceutical research, 1997

Interaction of human IgE with its high affinity receptor (Fc epsilon RI) on mast cells and basoph... more Interaction of human IgE with its high affinity receptor (Fc epsilon RI) on mast cells and basophils is an important step for initiating IgE mediated immune responses. To characterize the IgE and Fc epsilon RI interaction, we investigated this interaction in terms of stoichiometry and binding affinity in solution. The binding of IgE and IgE Fc epsilon RI alpha chain, the extracellular portion of IgE high affinity receptor (sFc epsilon RI alpha) was compared with the binding of IgE and IgE immunoadhesin (Fc epsilon RI alpha-IgG). The interaction was characterized by analytical ultracentrifugation, size exclusion chromatography, light scattering and ELISA. We show that the sFc epsilon RI alpha is only able to bind to one IgE, while the immunoadhesin can bind to two IgE. The interaction between IgE and Fc epsilon RI is very strong. Both forms of soluble receptors have similar intrinsic binding affinity with IgE. Both soluble receptors (Fc epsilon RI alpha-IgG and sFc epsilon RI alpha) ...

Pharmaceutical research, 1994

Recombinant human deoxyribonuclease I (rhDNase) is a new therapeutic agent developed to improve c... more Recombinant human deoxyribonuclease I (rhDNase) is a new therapeutic agent developed to improve clearance of purulent sputum from the human airways. It is delivered by inhalation. Four jet nebulizers, T Up-Draft II (Hudson), Customized Respirgard II (Marquest), Acorn II (Marquest), and Airlife Misty (Baxter), were evaluated in vitro for their ability to deliver aerosols of rhDNase. The aerosols were generated from 2.5-mL aqueous solutions of rhDNase, at concentrations of either 1 or 4 mg/mL. In all experiments, the Pulmo-Aide Compressor (De Vilbiss) was used to supply the air to the nebulizers. Between 20 and 28% of the rhDNase dose initially placed in the nebulizers was delivered to the mouthpiece in the respirable range (1-6 microns). Evaluation of the rhDNase following nebulization in all four devices indicated that there was no loss in enzymatic activity and no increase in aggregation. Circular dichroism spectrophotometry indicated there was no change in either the secondary or ...

Pharmaceutical biotechnology, 1993

Plasminogen activators (PA) are endogenous serine proteases involved in a cascade of events leadi... more Plasminogen activators (PA) are endogenous serine proteases involved in a cascade of events leading to the dissolution of a blood clot. These proteins are classified in two distinct groups: urokinase-type plasminogen activator (u-PA) and tissue-type plasminogen ...

Jameel/Manufacturing Biopharmaceuticals, 2010

... techniques, such as lyophilization and spray drying [6]. Other techniques, such as crystalliz... more ... techniques, such as lyophilization and spray drying [6]. Other techniques, such as crystallization [11] and microparticle-based technology [12 ... First, the rapid pumping and continuous circulation through narrow pathways may generate sufficient shear and cavitation stresses on a ...

Mahler/Protein Aggregates Analysis, 2012

ABSTRACT This chapter contains sections titled: Introduction Size Exclusion Chromatography Analyt... more ABSTRACT This chapter contains sections titled: Introduction Size Exclusion Chromatography Analytical Ultracentrifugation Field-Flow Fractionation Electrophoresis Other Potential Technologies Summary References

Journal of Chromatography B: Biomedical Sciences and Applications, 2001

Biochemistry, 1992

An expression plasmid encoding the extracellular portion of the human tumor necrosis factor (TNF)... more An expression plasmid encoding the extracellular portion of the human tumor necrosis factor (TNF) type 1 receptor (TNF-R1) was constructed and used to generate a stable cell line secreting soluble TNF-R1 (sTNF-R1). The sTNF-R1 was purified, and its biochemical properties and its interactions with human TNF-alpha were examined. SDS-PAGE resolved the purified sTNF-R1 into three bands of approximate Mr 24,200, 28,200, and 32,800. Sedimentation equilibrium analysis gave a molecular weight of 25,000 for sTNF-R1 whereas the molecular weight obtained by gel filtration chromatography was approximately 55,000-60,000. Scatchard analysis of [125I]TNF-alpha binding to sTNF-R1 revealed high-affinity binding (Kd = 93 pM), comparable to that observed for the intact receptor on whole cells. Competitive binding experiments showed that sTNF-R1 has a 50-60-fold higher affinity for TNF-alpha than for TNF-beta, in contrast to the equal affinities of TNF-alpha and TNF-beta for the full-length TNF-R1 transiently expressed in mammalian cells. The sTNF-R1 was found to block the cytotoxicity of TNF-alpha and TNF-beta on a murine L-M cell assay. The sizes of the sTNF-R1.TNF-alpha complex determined by gel filtration chromatography and sedimentation equilibrium were approximately 141 and 115 kDa, respectively. The stoichiometry of the complex was examined by Scatchard analysis, size-exclusion chromatography, HPLC separation, amino acid composition, sequence analysis, and sedimentation equilibrium. The data from these studies suggest that at least two molecules of sTNF-R1 can bind to a single TNF-alpha trimer.(ABSTRACT TRUNCATED AT 250 WORDS)

CRC critical reviews in biochemistry

Recent improvements in the understanding of electrostatic interactions in proteins serve as a foc... more Recent improvements in the understanding of electrostatic interactions in proteins serve as a focus for the general topic of pH-dependent processes in proteins. The general importance of pH-dependent processes is first set out in terms of hydrogen ion equilibria, stability, ligand interactions, assembly, dynamics, and events in related molecular systems. The development of various theoretical treatments includes various formalisms in addition to the solvent interface model developed by Shire et al. as an extension of the Tanford-Kirkwood treatment. A number of detailed applications of the model are presented and future potentialities are sketched.

... Auteur(s) / Author(s). CIPOLLA DC ; CLARK AR ; CHAN H.-K. ; GONDA I. ; SHIRE SJ ; Affiliation... more ... Auteur(s) / Author(s). CIPOLLA DC ; CLARK AR ; CHAN H.-K. ; GONDA I. ; SHIRE SJ ; Affiliation(s) du ou des auteurs / Author(s) Affiliation(s). Genentech Inc., South San Francisco CA 94080, ETATS-UNIS ... Langue / Language. Anglais Revue : Français Editeur / Publisher. ...

Journal of Biological Chemistry

A filter binding assay was developed to study interactions between purified TRAP, the trp RNA-bin... more A filter binding assay was developed to study interactions between purified TRAP, the trp RNA-binding attenuation protein of Bacillus subtilis, and trp specific transcripts. TRAP formed stable complexes with trpEDCFBA leader RNA; binding was L-tryptophan-dependent and was complete within 60 s. TRAP binds to a segment of the trp leader transcript that includes part of an RNA antiterminator structure. Binding to this segment allows formation of an RNA terminator structure, thereby promoting transcription termination. Using several trpEDCFBA leader deletion transcripts, we identified several closely spaced trinucleotide repeats (seven GAG and four UAG repeats) in the trp leader transcript that appeared to be required for TRAP binding. We also showed that TRAP binds to a segment of the trpG transcript that includes the trpG ribosome binding site; the nucleotide sequence of this segment contains several appropriately spaced trinucleotide repeats (seven GAG, one UAG, and one AAG). TRAP binding to the trpG transcript would block translation initiation. RNA footprint analysis confirmed interaction between TRAP and the trinucleotide repeats in the various transcripts. TRAP, in the presence or absence of L-tryptophan, appears to consist of 11 or 12 identical 8-kDa subunits. Our findings suggest that each tryptophan-activated TRAP subunit can bind one G/UAG repeat in a target transcript. Multiple protein-RNA interactions are required for stable association.

Ultrasensitive Biochemical Diagnostics II, 1997

ABSTRACT The use of analytical ultracentrifugation for probing macromolecular interactions in rec... more ABSTRACT The use of analytical ultracentrifugation for probing macromolecular interactions in recombinant deoxyribonucleic acid (DNA) technology was discussed. Analytical ultracentrifugation was used to determine absolute molecular weights without the use of molecular weight standards or interference from the sieving matrix used for separation. The instrument used for analytical ultracentrifugation comprised of an ultracentrifuge, designed to centrifuge components in a well controlled environment with good temperature control and a stable centrifugal field.

Monoclonal antibodies (MAb) have become a crucial therapeutic agent. However, some applications s... more Monoclonal antibodies (MAb) have become a crucial therapeutic agent. However, some applications such as a subcutaneous injection need highly concentrated protein solutions that can have undesired large viscosity. Therefore, it is of great importance to examine the relation between MAb self-associated nanostructure and its viscosity under various concentration and excipient conditions with techniques which can probe both structure and dynamics that span the length scale of an individual protein to a larger aggregate. We have investigated the protein-protein interactions and diffusive properties of highly concentrated MAbs using small angle neutron scattering (SANS) and neutron spin echo (NSE). Our results indicate that concentration, temperature, pH and surfactant do not have a strong effect on the individual MAb conformation for our MAb samples. The short-range attraction of some MAb proteins is found to be highly anisotropic in contrast to many other protein solutions at low ionic ...

The journal of physical chemistry. B, Jan 14, 2010

Light scattering intensity measurements of solutions of two purified monoclonal antibodies were p... more Light scattering intensity measurements of solutions of two purified monoclonal antibodies were performed over a wide range of concentrations (0.5-275 mg/mL) and ionic strengths (0.02 to 0.6 M). Despite extensive sequence homology between these mAbs, alteration of ∼20 amino acids in the complementarity determining regions resulted in different net intermolecular interactions and responses to solution ionic strength. The concentration dependence of scattering was analyzed by comparison with the predictions of three models, allowing for intermolecular interaction of various types. In order of increasing complexity, the three models account for: (1) steric repulsions (simple hard-sphere model), (2) steric repulsion with short-ranged attractive interactions of varying magnitude (adhesive hard-sphere model), and (3) steric and nonsteric repulsive interactions between several species whose relative concentrations may change as a function of total protein concentration as dictated by equil...

Pharmaceutical research, 2001

To determine the effect of moisture and the role of the glass transition temperature (Tg) on the ... more To determine the effect of moisture and the role of the glass transition temperature (Tg) on the stability of a high concentration, lyophilized, monoclonal antibody. A humanized monoclonal antibody was lyophilized in a sucrose/histidine/polysorbate 20 formulation. Residual moistures were from 1 to 8%. Tg values were measured by modulated DSC. Vials were stored at temperatures from 5 to 50 degrees C for 6 or 12 months. Aggregation was monitored by size exclusion chromatography and Asp isomerization by hydrophobic interaction chromatography. Changes in secondary structure were monitored by Fourier transform infrared (FTIR). T. values varied from 80 degrees C at 1% moisture to 25 degrees C at 8% moisture, there was no cake collapse and were no differences in the secondary structure by FTIR. All formulations were stable at 5 degrees C. High moisture cakes had higher aggregation rates than drier samples if stored above their Tg values. Intermediate moisture vials were more stable to aggr...

Pharmaceutical research, 1999

The purpose of this work is to utilize electron paramagnetic resonance (EPR) spectroscopy in conj... more The purpose of this work is to utilize electron paramagnetic resonance (EPR) spectroscopy in conjunction with analytical ultracentrifugation (AUC) to investigate the binding of surfactants to proteins with a transmembrance domain. As an example these methods have been used to study the interaction of a nonionic surfactant, C12E8, to recombinant human tissue factor (rhTF) in liquid formulations. The complementary nature of the two techniques aids in data interpretation when there is ambiguity using a single technique. In addition to binding stoichiometries, the possibility of identifying the interacting domains by using two forms of rhTF is explored. Two recombinant, truncated forms of human tissue factor were formulated in the absence of phospholipids. Neither of the recombinant proteins, produced in E. coli, contains the cytoplasmic domain. Recombinant human tissue factor 243 (rhTF 243) consists of 243 amino acids and includes the transmembrane sequences. Recombinant human tissue f...

Pharmaceutical research, 1999

To study the effect of trehalose, lactose, and mannitol on the biochemical stability and aerosol ... more To study the effect of trehalose, lactose, and mannitol on the biochemical stability and aerosol performance of spray-dried powders of an anti-IgE humanized monoclonal antibody. Protein aggregation of spray-dried powders stored at various temperature and relative humidity conditions was assayed by size exclusion chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis. Protein glycation was determined by isoelectric focusing and affinity chromatography. Crystallization was examined by X-ray powder diffraction. Aerosol performance was assessed as the fine particle fraction (FPF) of the powders blended with coarse carrier lactose, and was determined using a multiple stage liquid impinger. Soluble protein aggregation consisting of non-covalent and disulfide-linked covalent dimers and trimers occurred during storage. Aggregate was minimized by formulation with trehalose at or above a molar ratio in the range of 300: 1 to 500:1 (excipient:protein). However, the powder...

Pharmaceutical research, 1999

To develop a new technique, spray freeze drying, for preparing protein aerosol powders. Also, to ... more To develop a new technique, spray freeze drying, for preparing protein aerosol powders. Also, to compare the spray freeze-dried powders with spray-dried powders in terms of physical properties and aerosol performance. Protein powders were characterized using particle size analysis, thermogravimetric analysis, scanning electron microscopy, X-ray powder diffractometry, and specific surface area measurement. Aerosol performance of the powders was evaluated after blending with lactose carriers using a multi-stage liquid impinger or an Anderson cascade impactor. Two recombinant therapeutic proteins currently used for treating respiratory tract-related diseases, deoxyribonuclase (rhDNase) and anti-IgE monoclonal antibody (anti-IgE MAb), were employed and formulated with different carbohydrate excipients. Through the same atomization but the different drying process, spray drying (SD) produced small (approximately 3 microns), dense particles, but SFD resulted in large (approximately 8-10 m...

Pharmaceutical research, 1997

Interaction of human IgE with its high affinity receptor (Fc epsilon RI) on mast cells and basoph... more Interaction of human IgE with its high affinity receptor (Fc epsilon RI) on mast cells and basophils is an important step for initiating IgE mediated immune responses. To characterize the IgE and Fc epsilon RI interaction, we investigated this interaction in terms of stoichiometry and binding affinity in solution. The binding of IgE and IgE Fc epsilon RI alpha chain, the extracellular portion of IgE high affinity receptor (sFc epsilon RI alpha) was compared with the binding of IgE and IgE immunoadhesin (Fc epsilon RI alpha-IgG). The interaction was characterized by analytical ultracentrifugation, size exclusion chromatography, light scattering and ELISA. We show that the sFc epsilon RI alpha is only able to bind to one IgE, while the immunoadhesin can bind to two IgE. The interaction between IgE and Fc epsilon RI is very strong. Both forms of soluble receptors have similar intrinsic binding affinity with IgE. Both soluble receptors (Fc epsilon RI alpha-IgG and sFc epsilon RI alpha) ...

Pharmaceutical research, 1994

Recombinant human deoxyribonuclease I (rhDNase) is a new therapeutic agent developed to improve c... more Recombinant human deoxyribonuclease I (rhDNase) is a new therapeutic agent developed to improve clearance of purulent sputum from the human airways. It is delivered by inhalation. Four jet nebulizers, T Up-Draft II (Hudson), Customized Respirgard II (Marquest), Acorn II (Marquest), and Airlife Misty (Baxter), were evaluated in vitro for their ability to deliver aerosols of rhDNase. The aerosols were generated from 2.5-mL aqueous solutions of rhDNase, at concentrations of either 1 or 4 mg/mL. In all experiments, the Pulmo-Aide Compressor (De Vilbiss) was used to supply the air to the nebulizers. Between 20 and 28% of the rhDNase dose initially placed in the nebulizers was delivered to the mouthpiece in the respirable range (1-6 microns). Evaluation of the rhDNase following nebulization in all four devices indicated that there was no loss in enzymatic activity and no increase in aggregation. Circular dichroism spectrophotometry indicated there was no change in either the secondary or ...

Pharmaceutical biotechnology, 1993

Plasminogen activators (PA) are endogenous serine proteases involved in a cascade of events leadi... more Plasminogen activators (PA) are endogenous serine proteases involved in a cascade of events leading to the dissolution of a blood clot. These proteins are classified in two distinct groups: urokinase-type plasminogen activator (u-PA) and tissue-type plasminogen ...

Jameel/Manufacturing Biopharmaceuticals, 2010

... techniques, such as lyophilization and spray drying [6]. Other techniques, such as crystalliz... more ... techniques, such as lyophilization and spray drying [6]. Other techniques, such as crystallization [11] and microparticle-based technology [12 ... First, the rapid pumping and continuous circulation through narrow pathways may generate sufficient shear and cavitation stresses on a ...

Mahler/Protein Aggregates Analysis, 2012

ABSTRACT This chapter contains sections titled: Introduction Size Exclusion Chromatography Analyt... more ABSTRACT This chapter contains sections titled: Introduction Size Exclusion Chromatography Analytical Ultracentrifugation Field-Flow Fractionation Electrophoresis Other Potential Technologies Summary References

Journal of Chromatography B: Biomedical Sciences and Applications, 2001