Platelet function testing: state of the art.

Platelet function testing has evolved from crude tests, such as the bleeding time, to tests that permit a relatively sophisticated evaluation of platelet activity. Nonetheless, these tests are hampered by lack of specificity and sensitivity, and poor standardization of methods and techniques. The bleeding time, which has long been a staple of hemostasis testing, has been dropped from the test menu at many laboratories.

Diagnosis and management of heparin-induced thrombocytopenia.

Heparin use is ubiquitous, wherein 1 to 5% of patients exposed to standard unfractionated heparin develop thrombocytopenia due to antibodies to a complex of heparin and platelet factor 4. Classic features include onset of thrombocytopenia after 5 to 10 days of ongoing heparin exposure, a 50% fall in the platelet count from baseline, resolution of the thrombocytopenia 5 to 10 days after cessation of heparin and a high risk of thrombosis noted in 30 to 75% of patients with heparin-induced thrombocytopenia (HIT) in terms of every-other-day platelet-count monitoring in patients on standard unfractionated heparin. And those patients developing thrombocytopenia necessitate an accurate, readily accessible diagnostic test for HIT.

Substrate specificity of the Saccharomyces cerevisiae Mus81-Mms4 endonuclease.

Mus81-Mms4/Eme1 is a conserved structure-specific endonuclease that functions in mitotic and meiotic recombination. It has been difficult to identify a single preferred substrate of this nuclease because it is active on a variety of DNA structures. In addition, it has been suggested that the specificity of the recombinant protein may differ from that of the native enzyme.

Slx1-Slx4 is a second structure-specific endonuclease functionally redundant with Sgs1-Top3.

The RecQ DNA helicases human BLM and yeast Sgs1 interact with DNA topoisomerase III and are thought to act on stalled replication forks to maintain genome stability. To gain insight into this mechanism, we previously identified SLX1 and SLX4 as genes that are required for viability and for completion of rDNA replication in the absence of SGS1-TOP3. Here we show that SLX1 and SLX4 encode a heteromeric structure-specific endonuclease.

The mechanism of Mus81-Mms4 cleavage site selection distinguishes it from the homologous endonuclease Rad1-Rad10.

Mus81-Mms4 and Rad1-Rad10 are homologous structure-specific endonucleases that cleave 3' branches from distinct substrates and are required for replication fork stability and nucleotide excision repair, respectively, in the yeast Saccharomyces cerevisiae. We explored the basis of this biochemical and genetic specificity. The Mus81-Mms4 cleavage site, a nick 5 nucleotides (nt) 5' of the flap, is determined not by the branch point, like Rad1-Rad10, but by the 5' end of the DNA strand at the flap junction.