Fibrin degradation product (FnDP) assays: analysis of standardization issues and target antigens in plasma.

The in vivo formation of fibrin and its subsequent secondary fibrinolytic digestion yields a variety of crosslinked fibrin degradation products (XL/FnDP). One of these is known as D-dimer and a variety of ELISA-type commercial kits using monoclonal antibodies to D-dimer have been developed. We have investigated the possibility of establishing a standard such that these various kits might indicate the same levels of D-dimer in the same samples.

Generation and partial characterization of five monoclonal antibodies with high affinities for fibrin.

Five fibrin-specific monoclonal antibodies have been generated using three different immunogens and a unique clot/plasma screening procedure. Two of the mAbs are directed to an epitope(s) on the A alpha-chain of fibrinogen while two others seem to react with a distinct B beta-chain epitope(s). A third antibody seems to displace all the mAbs from fibrin and may be directed to a repeating structural domain in the fibrin.

A comparison of the plasminogen activator activity units of urinary-type plasminogen activator (u-PA) and tissue-type plasminogen activator (t-PA).

The earliest tissue plasminogen activator (t-PA) preparations prepared from melanoma cells were expressed in urinary-type plasminogen activator (u-PA) units of activity using the u-PA International Standard (66/46). This report describes a comparison between u-PA and t-PA units by various types of fibrin plate procedure using both human and bovine fibrin. Within the biometric limits of this procedure it was found that the potency ratio of u-PA/t-PA was 3.

Screening for fibrin specific monoclonal antibodies: the development of a new procedure.

The acquisition of monoclonal antibodies specific for human fibrin has been impaired by the similarity in chemical composition between fibrinogen and fibrin and the conformational difference between immobilised and soluble fibrinogen. Five monoclonal antibodies (mabs) with a known affinity for fibrin have been subjected to screening procedures which involved the presentation of different forms of both fibrinogen and fibrin to the test mabs. It was observed by scanning electron microscopy that dried fibrin (denoted fibrin D), immobilised on the wells of PVC plates was morphologically similar to the fibrin found in human clots whereas PVC-immobilised fibrin monolayers (fibrin M) and a homogenised form of fibrin (fibrin FF) presented two very different morphological appearances.

A novel sialylated N-acetylgalactosamine-containing oligosaccharide is the major complex-type structure present in Bowes melanoma tissue plasminogen activator.

We have employed fast atom bombardment mass spectrometry (FAB-MS) to screen the N-linked oligosaccharides of Bowes melanoma tissue plasminogen activator (mt-PA), and recombinant t-PAs produced by Chinese hamster ovary cells (rt-PA) and by a gene-enriched melanoma cell line (rmt-PA). These studies have confirmed the published structures for rt-PA, but are not in agreement with some of the structures reported for mt-PA. In the latter glycoprotein we have identified a novel structure as the major oligosaccharide attached to Asn-184 and Asn-448.