Jerry B. Lefkowitz

The prothrombin Denver patient has two different prothrombin point mutations resulting in Glu‐300→Lys and Glu‐309→Lys substitutions

Dysprothrombinaemia is a rare, congenital cause of bleeding. Fewer than 25 families who express a functional prothrombin (factor II) defect have been reported. The original patient with prothrombin Denver had a severe haemophilia‐like bleeding disorder treated with weekly prophylactic factor replacement. Analysis of factor II activity and antigen in the patient showed a factor II activity of 5 units/dl and factor II antigen of 21 units/dl. Genomic DNA from the patient, mother and brother was obtained from peripheral blood white cells. Oligonucleotides were constructed, and prothrombin exons were amplified via polymerase chain reaction (PCR). The entire sequence of the thrombin portion of the molecule (exons VIII–XIV) and that of exons I–II and IV–VII was determined. This moderately severe dysprothrombinaemia was found to be associated with compound heterozygosity for two different Glu→Lys point mutations, at amino acid positions 300 and 309. Assays of plasma from the prothrombin Denver proband suggested that the functional defect was in the activation of zymogen to enzyme.

An institution-specific heparin titration nomogram: development, validation, and assessment of compliance.

Study Objective. To develop, validate, and assess compliance with a heparin titration nomogram.

Fibrinogen Longmont. A heterozygous abnormal fibrinogen with B beta Arg-166 to Cys substitution associated with defective fibrin polymerization.

Abstract: BβArg166 to Cys substitution was identified in an abnormal fibrinogen named fibrinogen Longmont. The proband, a young woman, and her mother were heterozygous; both experienced episodes of severe hemorrhage at childbirth. The neo‐Cys residues were found to be disulfide‐bridged to either an isolated Cys amino acid or to the corresponding Cys residue of another abnormal fibrinogen molecule, forming dimers. Thrombin and batroxobin induced fibrin polymerization were impaired, despite normal release of fibrinopeptides A and B. Moreover, the polymerization defect was not corrected by removing the dimeric species or adding calcium. Fibrinogen Longmont had normal polymerization site a, as evidenced by normal GPRP‐peptide binding. Thus, the sites A and a can interact to form protofibrils, as evidenced by dynamic light scattering measurements. These protofibrils, however, do not associate laterally in a normal manner, leading to an abnormal clot formation.

Argatroban-coupled Affi-Gel matrix for the purification of thrombin from plasma

Sometimes it is necessary to obtain thrombin from limited amounts of human plasma for laboratory assay. None of the available purification methods easily deals with this subject. The procedure described in the present paper uses a readily available pharmaceutical agent, argatroban, to construct an affinity matrix. Argatroban has a high affinity for thrombin and its thrombin binding is reversible. Prothrombin derived from a Ba2+ precipitate of human plasma is used as the starting material. The crude prothrombin can be bulk activated to thrombin using taipan‐snake (Oxyuranus scutellatus) venom and bound to the argatroban‐coupled matrix without further processing steps. The thrombin product eluted from the argatroban matrix is very pure as judged by high specific activity and by electrophoresis. This purification scheme is rapid, yielding purified thrombin within 2 days.