scholarly journals Rapid quantitative analysis of monoclonal antibody heavy and light chain charge heterogeneity

mAbs ◽  
2015 ◽  
Vol 7 (6) ◽  
pp. 1118-1127 ◽  
Author(s):  
Ram P Vanam ◽  
Michael A Schneider ◽  
Michael S Marlow
Keyword(s):  
Monoclonal Antibody ◽  
Quantitative Analysis ◽  
Light Chain ◽  
Charge Heterogeneity

Characterization of Monocyte-Associated Factor V

1993 ◽  
Vol 70 (02) ◽  
pp. 273-280 ◽  
Author(s):  
Janos Kappelmayer ◽  
Satya P Kunapuli ◽  
Edward G Wyshock ◽  
Robert W Colman
Keyword(s):  
Monoclonal Antibody ◽  
Light Chain ◽  
Peripheral Blood ◽  
De Novo ◽  
Factor V ◽  
Blood Monocytes ◽  
De Novo Synthesis ◽  
Hep G2 ◽  
Hep G2 Cells ◽  
Peripheral Blood Monocytes

SummaryWe demonstrate that in addition to possessing binding sites for intact factor V (FV), unstimulated peripheral blood monocytes also express activated factor V (FVa) on their surfaces. FVa was identified on the monocyte surface by monoclonal antibody B38 recognizing FVa light chain and by human oligoclonal antibodies H1 (to FVa light chain) and H2 (to FVa heavy chain) using immunofluorescence microscopy and flow cytometry. On Western blots, partially cleaved FV could be identified as a 220 kDa band in lysates of monocytes. In addition to surface expression of FVa, monocytes also contain intracellular FV as detected only after permeabilization by Triton X-100 by monoclonal antibody B10 directed specifically to the Cl domain not present in FVa. We sought to determine whether the presence of FV in peripheral blood monocytes is a result of de novo synthesis.Using in situ hybridization, no FV mRNA could be detected in monocytes, while in parallel control studies, factor V mRNA was detectable in Hep G2 cells and CD18 mRNA in monocytes. In addition, using reverse transcriptase and the polymerase chain reaction, no FV mRNA was detected in mononuclear cells or in U937 cells, but mRNA for factor V was present in Hep G2 cells using the same techniques. These data suggest that FV is present in human monocytes, presumably acquired by binding of plasma FV, and that the presence of this critical coagulation factor is not due to de novo synthesis.

Purification of Urokinase from Complex Mixtures Using Immobilized Monoclonal Antibody Against Urokinase Light Chain

1983 ◽  
Vol 49 (01) ◽  
pp. 024-027 ◽  
Author(s):  
David Vetterlein ◽  
Gary J Calton
Keyword(s):  
Monoclonal Antibody ◽  
Light Chain ◽  
Culture Media ◽  
Biological Fluids ◽  
Single Step ◽  
High Yield ◽  
Step Method ◽  
Commercial Preparation ◽  
E Coli ◽  
Tissue Culture Media

SummaryThe preparation of a monoclonal antibody (MAB) against high molecular weight (HMW) urokinase light chain (20,000 Mr) is described. This MAB was immobilized and the resulting immunosorbent was used to isolate urokinase starting with an impure commercial preparation, fresh urine, spent tissue culture media, or E. coli broth without preliminary dialysis or concentration steps. Monospecific antibodies appear to provide a rapid single step method of purifying urokinase, in high yield, from a variety of biological fluids.

A Factor VIII Neutralizing Monoclonal Antibody and a Human Inhibitor Alloantibody Recognizing Epitopes in the C2 Domain Inhibit Factor VIII Binding to von Willebrand Factor and to Phosphatidylserine

1993 ◽  
Vol 69 (03) ◽  
pp. 240-246 ◽  
Author(s):  
Midori Shima ◽  
Dorothea Scandella ◽  
Akira Yoshioka ◽  
Hiroaki Nakai ◽  
Ichiro Tanaka ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Amino Acid ◽  
Factor Viii ◽  
Von Willebrand Factor ◽  
Light Chain ◽  
Amino Acid Residues ◽  
Amino Terminal ◽  
Neutralizing Monoclonal Antibody ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryA neutralizing monoclonal antibody, NMC-VIII/5, recognizing the 72 kDa thrombin-proteolytic fragment of factor VIII light chain was obtained. Binding of the antibody to immobilized factor VIII (FVIII) was completely blocked by a light chain-specific human alloantibody, TK, which inhibits FVIII activity. Immunoblotting analysis with a panel of recombinant protein fragments of the C2 domain deleted from the amino-terminal or the carboxy-terminal ends demonstrated binding of NMC-VIII/5 to an epitope located between amino acid residues 2170 and 2327. On the other hand, the epitope of the inhibitor alloantibody, TK, was localized to 64 amino acid residues from 2248 to 2312 using the same recombinant fragments. NMC-VIII/5 and TK inhibited FVIII binding to immobilized von Willebrand factor (vWF). The IC50 of NMC-VIII/5 for the inhibition of binding to vWF was 0.23 μg/ml for IgG and 0.2 μg/ml for F(ab)'2. This concentration was 100-fold lower than that of a monoclonal antibody NMC-VIII/10 which recognizes the amino acid residues 1675 to 1684 within the amino-terminal portion of the light chain. The IC50 of TK was 11 μg/ml by IgG and 6.3 μg/ml by F(ab)'2. Furthermore, NMC-VIII/5 and TK also inhibited FVIII binding to immobilized phosphatidylserine. The IC50 for inhibition of phospholipid binding of NMC-VIII/5 and TK (anti-FVIII inhibitor titer of 300 Bethesda units/mg of IgG) was 10 μg/ml.

A monoclonal antibody Hy20-54-16-3L to lambda (λ) light chain of human immunoglobulin reacts with amyloid in Alzheimer's disease brain

1990 ◽  
Vol 117 (1-2) ◽  
pp. 62-67 ◽  
Author(s):  
Fuyuki Kametani ◽  
Kikuko Tanaka ◽  
Maroto Sato ◽  
Seiichi Haga ◽  
Takashi Saitoh ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Monoclonal Antibody ◽  
Light Chain ◽  
Human Immunoglobulin ◽  
Λ Light Chain

Monoclonal Antibody Light Chain with Prothrombinase Activity†

Biochemistry ◽  
2000 ◽  
Vol 39 (21) ◽  
pp. 6459-6465 ◽  
Author(s):  
Perumal Thiagarajan ◽  
Robert Dannenbring ◽  
Kinji Matsuura ◽  
Alfonso Tramontano ◽  
Gennady Gololobov ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Light Chain ◽  
Prothrombinase Activity

Preparation and Immunological Traits of Monoclonal Antibody against Sarafloxacin

2012 ◽  
Vol 459 ◽  
pp. 54-57
Author(s):  
Guo Ying Fan ◽  
Jin Qing Jiang
Keyword(s):  
Monoclonal Antibody ◽  
Detection Limit ◽  
Cell Fusion ◽  
Light Chain ◽  
Dynamic Range ◽  
Fusion Technology ◽  
Alternative Method ◽  
Ic50 Values ◽  
The Stability ◽  
Igg1 Isotype

Through cell fusion technology, five hybridoma lines of sarafloxacin (SAR), named S1-B2, S2-C6, S2-E7, S3-C5, and S3-E5, were identified and their corresponding mAbs were of the IgG1 isotype with a k light chain. The Kaffs of all mAbs were between 2.8 and 4.6×109 L/mol. The titers and IC50 values of purified ascite fluids were in the range of 0.512–2.56×106 and 0.32–0.48 ng/mL, respectively. The performances of S1-B2 and S2-C6 were more consistent in the stability experiments. Based on the S1-B2 hybridoma, an icELISA method was developed. The dynamic range was from 0.004 to 18 ng/mL, with a detection limit for the assay and IC50 values of 0.002 and 0.32 ng/mL, respectively. Therefore, the establishment of these hybridomas may provide an alternative method for the detection of SAR residues in food-original animals.

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