Single Step Purification of Platelet Factor XIII Using an Immobilized Factor XIII A-Subunit Monoclonal Antibody

Lukácová, D; Reed, Guy L.

doi:10.1055/s-0038-1651620

Cited by 5 publications

(2 citation statements)

References 23 publications

(27 reference statements)

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“…To obtain immunoinhibitors of FXIII, we initially immunized mice with A2B2. Somatic cell fusion with splenocytes from these mice yielded many anti-FXIII MAbs but only one was directed against A2 (29). We then immunized mice with A2 alone.…”

Section: Resultsmentioning

confidence: 99%

Generation and Mechanism of Action of a Potent Inhibitor of Factor XIII Function

Reed

Lukácová

1995

Thromb Haemost

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SummaryAlthough known to play a critical role in thrombosis, the precise role of factor XIII in other processes like wound healing and gestation remains to be elucidated. Because a specific, potent inhibitor could help define the function of factor XIII in these processes, or determine the potential benefits of factor XIII suppression in thrombotic disease, we have derived and characterized a monoclonal antibody inhibitor of factor XIII activation. This immunoinhibitor, 9C11, reacts specifically with both plasma and platelet forms of factor XIII. When added to human plasma, either as whole immunoglobulin or as Fab fragments, 9C11 completely prevented thrombin-activated factor XIII activity with small molecular weight substrates like 14C-putrescine. In addition, when clotted with plasma, 9C11 ablated the gamma chain crosslinking of fibrin catalyzed by factor XIII and markedly accelerated the fibrinolysis of plasma clots by urokinase. Studies of the mechanism of action showed that 9C11 inhibited the cleavage and activation of factor XIII by thrombin, but did not affect the catalytic function of previously activated factor XIII. Although it inhibited thrombin activation, experiments indicated that it bound comparably to both factor XIII zymogen and the thrombin-cleaved zymogen, and did not bind to the thrombin cleavage site of the molecule. Taken together these experiments indicated that 9C11 acted as an inhibitor of the thrombin cleavage and activation of the factor XIII A-subunit. Its potency and specificity make it a useful agent for studying thrombin-activatable factor XIII function in biological systems.

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Section: Resultsmentioning

confidence: 99%

Generation and Mechanism of Action of a Potent Inhibitor of Factor XIII Function

Reed

Lukácová

1995

Thromb Haemost

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“…As a common experience, it has been difficult to make m()llocl()nal antibodies against native. factor XIII but recently Lukacova and Reed (35) have reported isolation of one antibody, caned 6D6, that they used to immunopurify platelet factor XIII. Onthe other hand, polyclonal antisera that inhibit factor XIII, as does .R-14, have been reported and a few are commercially available.…”

Section: Discussionmentioning

confidence: 99%

Characterization of a Monoclonal Antibody Directed Against the Carboxyl-Terminus of Human Factor XIII

et al. 1994

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SummaryBy deriving an anti-peptide monoclonal antibody, mAb 7A4, we characterized the relatively unstudied carboxyl-terminal end of the α-chain of human factor XIII, the plasma transglutaminase. MAb 7A4 was directed against the last eight amino acids (Gln-Ile-Gln-Arg-Arg-Pro-Ser-Met) and bound with a dissociation constant of 3.4 × 10−8 M. In a solid assay format, mAb 7A4 bound equally well to factor XIII obtained from human plasma, platelets or placenta. However, in a solution-phase assay format, the epitope was largely unavailable but could be readily exposed by heat denaturation. Tmmunoblotting showed that this epitope is conserved among all species of plasma factor XIII tested except rabbit suggesting that the carboxyl-terminus might be an important structural element. Other competitive binding experiments with synthetic peptides as inhibitors pointed toward the final carboxyl-terminal amino acid, Met-731, as an immunochemically important determinant. This was used advantageously to confirm the finding that the caiboxyl-leiminal Mel-731 is largely absent from placental factor XIII (1) as compared to platelet or plasma factor XIII.

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Transglutaminases: purification and actuivity assays

Wilhelm

Meinhardt

Seitz

1996

Journal of Chromatography B: Biomedical Sciences and Applicatio

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Single Step Purification of Platelet Factor XIII Using an Immobilized Factor XIII A-Subunit Monoclonal Antibody

Cited by 5 publications

References 23 publications

Generation and Mechanism of Action of a Potent Inhibitor of Factor XIII Function

Generation and Mechanism of Action of a Potent Inhibitor of Factor XIII Function

Characterization of a Monoclonal Antibody Directed Against the Carboxyl-Terminus of Human Factor XIII

Transglutaminases: purification and actuivity assays

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