Partial Purification of ADAMTS-13

POWAZNIAK, YANINA; KEMPFER ANA C; AMARAL MARÍA M; FARÍAS, CRISTINA; VERA, EZEQUIEL; LAZZARI MARÍA A

Sydney

Congreso; XX CONGRESS. THE INTERNATIONAL SOCIETY ON THROMBOSIS AND HAEMOSTASIS AND 51st ANNUAL SSC METTING; 2005

THE INTERNATIONAL SOCIETY ON THROMBOSIS AND HAEMOSTASIS

We developed the partial purification (P) of ADAMTS-13 by methods (M) in which cryoprecipitates were prepared from: M1- plasma, M2- plasma treated with polyetilenglicol (PEG) 4% (known M) and M3- supernatant obtained from M1 treated with PEG 4%, M4- dialyzed precipitate (dissolved in saline) obtained from M2 (combined M). Assays: VWF:Ag , ADAMTS-13, fibrinogen (Fg) (100U/dL= 390 mg/dL assayed by Clauss M), proteins. P was expressed as specific activity (SA) of samples (U/g of protein)/ /SA of the original plasma (14 U/g). P results (mean ±SD, n=5) of precipitate (p) and supernatant (s) fractions (Fr) obtained by centrifugation from M1 to M4 are in the Table. Fr ADAMTS-13 VWF:Ag ADAMTS-13/VWF:Ag Fg ADAMTS-13/Fg 1p 1,4 ±0,6 45±20 0.03 23±10 0.06 1s 1.5±0.8 0,2±0,1 7.50* 0,4±0.2 3.75* 2p 11±5 * 20±10 0.55 17±12 0.65 2s 1,0±0.3 0,4±0,2 2.50* 0,6±0,4 1.67 3p 7±4 * 4±2 1.75* 3.5±0.7 2.00* 3s 1.5±0,5 0,3±0,2 5.00* 0.6±0.3 2.67* 4p 16±8* 33±18 0.48 16±12 1.00 4s 6±2* 3±2 2.00* 13±3 0.50 The Fr 3p fulfills the three requirements (adequate P and not so high contamination of VWF:Ag and Fg) considering 1.75 (with asterisk) as cut-off level. This Fr could be used like source to purify ADAMTS-13. Treatment options for thrombotic thrombocytopenic purpura include fresh frozen plasma infusion. Aronson D (VS 2003), assaying Cohn Fr, suggested that the use of partially purified Fr may be particularly valuable in lowering the volume administeredand decreasing the risk of allergic reactions. The therapeutic use of these new Fr from combined M must be investigated.