A question about pooled normal plasma for lupus anticoagulant testing from Theo Powers, US Oncology.
Hello, George! Great web site,
The updated guidelines for lupus anticoagulant (LA) testing refer to making in-house pooled normal plasma (PNP). What are the current recommendations for preparing this pool, such as number of donors, storage, stability? Triplett (Laboratory Evaluation of Coagulation, 1982) recommended six normal donors. I’ve seen recommendations for 10, 20, 40 normal donors. I look forward to hearing your reply, as my resources are older. Theo Powers MT (ASCP)
Hi, Theo, thank you for your question. To avoid the appearance of a Precision BioLogicbias, I ran this past my friend and frequent contributor, Dave McGlasson at Wilford Hall USAF Hospital in San Antonio. Here’s Dave’s response:
George,
According to Pengo V, et al: Update of the guidelines for lupus anticoagulant detection. J Thromb Haemostas 2009;7:1737-40 and Tripodi A: Laboratory testing for lupus anticoagulants: Diagnostic criteria and use of screening, mixing, and confirmatory studies. Sem Thromb Hemost 2008;34:373-9, the following summary guidelines should be adhered to when preparing PNP:
1. Donor plasmas should have normal levels, close to 100% activity, for all the coagulation factors and should be stored at -70°C to preserve their activity.
2. Local normal plasmas may be prepared by pooling plasmas of 15 or more donations from healthy subjects free from hemorrhagic or thrombotic diseases who are not taking drugs known to impair hemostasis.
3. The material must be platelet-free and may be stored in small aliquots.
4. Commercial frozen normal plasmas can be used if they fulfill the above specifications or have otherwise been validated for use in LA testing.
Dave adds that in order to prepare PNP in-house you must perform PT, PTT, fibrinogen and all factor assays on donor plasmas before processing and storage. The normal plasma should “contain minimal residual platelets (<10,000/uL plasma).” Precision BioLogic’s CRYOcheck™ PNP contains plasma from a minimum of 20 healthy normal males and females negative for LA. Precision validates the plasma collection process to ensure platelet counts are less than 10,000/uL and provides a Certificate of Analysis to confirm all factors are present at near 100%. Laboratories collecting in-house pools will need to ensure this as well.
[Inserted by George: For safety, plasmas should also be confirmed negative for HBV, HCV, and HIV. Just so this doesn’t appear to be pure self-promotion, George King Biomedical also distributes high-quality frozen plasmas. All others distribute lyophilized materials.]
Although labs may suggest that local preparation doesn’t cost very much, it might be worthwhile to ask, “is there something more valuable I could be doing with my time?” Not to mention the actual cost of performing all of the assays necessary to confirm the true normal accuracy of your local PNP. Further, as labs lose staff and as staff age, there are fewer non-medicated normal donors to collect from.
David L. McGlasson, MS, MLS(ASCP)CM
59th Clinical Research Division
Wilford Hall Medical Center
Lackland AFB, TX
Thank you, George, for mentioning George King Bio-Medical, I
Thank you, George, for mentioning George King Bio-Medical, Inc. I would also like to add George King Bio-Medicals Pooled Normal Plasma consists of 30 or more carefully screened donors of which are non-medicated, nonsmoking, both male and female, from a diverse population. George King Bio-Medical Plasma meets guidelines required by CAP & CLSI. A laboratory would greatly benefit by using commercially prepared Pooled Normal Plasma versus making an in-house Pooled Normal Plasma.
Steve Duff of Precision BioLogic adds this helpful update:
Steve Duff of Precision BioLogic adds this helpful update:
Hi David:
Just noting your post on Fritsma Factor… well done! I would note that on your point 3. indicating plasmas must be “platelet-free”, I believe the guideline states… “The PNP should be prepared ad hoc (home-made) by double centrifugation to ensure that the PNP contains minimal residual platelets (<107/mL) ." Thank you, Steve, we sometimes get tangled up in definitions for platelet-poor and platelet-free. Geo.