PNP: How Many Donors?

PNP: How Many Donors?
George Fritsma
Mar 10, 2016 10:24am

From Dr. Tang: Dear George, In the mixing test, commonly 20 healthy donors is required for preparing normal pooled plasma, why not 15, 10, or 5 donors? Have any studies validated that? Thank you for your reply.

Hello, Dr. Tang. I know of no published studies that establish 20 donors as a minimum for producing pooled normal plasma. The policy appears to be based on custom and expert opinion. I've invited some experts to comment, we may learn more in the next few days. Geo.

More: I just spoke with Larry Brace, PhD, who asserts that 20 data points provide statistical assurance that the pool results represent a true population sampling, and that this is a statistician's convention. However, we don't have access to authoritative references.

Specimen Management, Validation
4 Comments

From Dr. Tang: Dear George, In the mixing test, commonly 20 healthy donors is required for preparing normal pooled plasma, why not 15, 10, or 5 donors? Have any studies validated that? Thank you for your reply.

Hello, Dr. Tang. I know of no published studies that establish 20 donors as a minimum for producing pooled normal plasma. The policy appears to be based on custom and expert opinion. I've invited some experts to comment, we may learn more in the next few days. Geo.

More: I just spoke with Larry Brace, PhD, who asserts that 20 data points provide statistical assurance that the pool results represent a true population sampling, and that this is a statistician's convention. However, we don't have access to authoritative references.

By Dr. Ali Sadeghi-khomami
Mar 10, 2016 3:03pm
The reason should stem from statistics and plasma component variations in human population. Considering the challenges that exist in defining a true “normal individual” for coagulation testing, after blending plasma from at least 20 individuals the chance of bias would be minimal. A few examples of interfering conditions in coagulation testing, which sometimes remain non-detected by routine screening criteria when searching for apparently healthy individuals, are: inflammation (FVIII and C-reactive protein elevation), special diet, age variation, single gender, taking medicines, mild infection, transient lupus-like antibodies, smoking, etc.
If the question is why 20 and not 15 plasma samples; well then it depends on how confident you want to be in your final result, and the trade-off you have to make between logistics and accuracy in your diagnosis. The following text is from Standard CLSI H47A2: The NPP should be prepared from a minimum of 20 donors or purchased from a commercial supplier, be cell-free (platelets may neutralize LAs), be fresh-frozen, possess ~100% factor levels, and have a PTT close to the mean of the reference range."
FYI, Precision BioLogic is offering Cryocheck Pooled Normal Plasma so that labs could make diagnosis with confidence without any need to go through the challenge associated with logistic of PNP preparation, running all required coagulation screening tests and reserve long lasting supply.
By Dr. Ning Tang
Mar 10, 2016 10:18pm
Thank you for the detailed answers. In China, commercial PNP is not available. in a recent web survey I initiated, about 80% of clinical labs in the whole of China have not carried out mixing tests, partly due to the difficulty in preparing home-made PNP. Whether a commercial normal control plasma can be used as PNP, if the targeted levels of each coagulation factor are stated about 100%?
From Geo, yes, the PNPs all have stated factor levels of approximately 100%. Thank you for your comment.
By Dr. Ali Sadeghi-khomami
Mar 11, 2016 1:04pm
Regarding Dr Tang’s recent question: “Whether a commercial normal control plasma can be used as PNP, if the targeted levels of each coagulation factor are stated about 100%?” Based on the normal control's intended use, the short answer is no. A normal control has different preparation requirements and it may not be pooled from several normal individuals as instructed in CLSI. Also, many commercial normal controls are lyophilized and adulterated with stabilizing agent and preservatives. Thus they could not be used safely in mixing study, especially for temperature dependent inhibitor detection which requires 2h incubation after mixing. Normal controls are not the same as pooled normal plasma thus their application in a mixing study is not recommended.
By Director of Marketing and Business Development Dean Willett
Mar 24, 2016 9:51am
Dear Dr. Tang, With all due respect to Dr. Sadeghi-Khomami, the ISTH SSC guidelines for LA testing do allow for the use of commercial lyophilized normal plasmas in mixing tests. I would recommend the following article. It could be useful:
http://onlinelibrary.wiley.com/doi/10.1111/j.1538-7836.2009.03555.x/full

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