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Gathering evidence for BCSH FMH guidelines ‐ counting controversies in FMH
Author(s) -
White J. L.,
Milkins C. E.,
Rowley M. R.,
Lubenko A.
Publication year - 2006
Publication title -
transfusion medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.471
H-Index - 59
eISSN - 1365-3148
pISSN - 0958-7578
DOI - 10.1111/j.1365-3148.2006.00694_27.x
Subject(s) - bleed , medicine , surgery
The BCSH guidelines for FMH first published in 1999 are now being updated. Evidence on which some of the recommendations are made is being re‐examined, e.g. the number of cells to be counted using the acid elution method (AE), to identify and quantify fetal bleeds. When using flow cytometry (FC) to measure FMH, the guidelines recommend that 500,000 events be collected, based on the statistics of rare event analysis where the CV% of the method decreases with the number of events collected (see Fig. 1). Extrapolating this statistical principle to acid elution (1 in 1000 ‘event’ equates to a 2.4ml bleed using Mollison's formula), where counting 6000 adult cells for FMH quantification is currently recommended, the CV% would be approximately 60 for a 4ml bleed. However this can be improved to approx. 25 if 10,000 cells are counted.1[ CV% ( y ‐axis) vs number of events counted ( x ‐axis) for a Poisson distribution where approx. 1 in 1000 positive events is expected. ] These statistics can also be applied to screening by FMH, where identification of bleeds >4ml is essential since the most commonly used post‐natal dose of anti‐D immunoglobulin (500 i.u.) covers a 4mL bleed. When scanning 25 low power fields (lpf) using x10 objective and x10 eyepiece, a minimum of 50,000 cells are screened (based on data from Mollison and UK NEQAS). The inter‐laboratory CV% = 38 for bleeds of 2 ‐ 6ml (by AE) in UK NEQAS exercises and the CV% according to rare event statistics is approximately 20. These figures can be used to predict the range of FMH values observed at different bleed volumes (see table), using the formula SD = (CV x mean) / 100 and where the expected bleed volume is the mean.mL FMH Expected range of FMH (mL) at 95% confidence (=2SD) CV% 20 (rare event) CV% = 38 (UK NEQAS)4 2.4–5.6 1.0–7.0 2 1.2–2.8 0.5–3.5Theoretically, 2ml is a safer trigger for quantification and follow‐up, since with a 4ml ‘cut‐off’ there is a risk that FMH of >4ml will not be recognised on screening 25 lpf, even where proficiency is good. Also, given the predicted high CV% for quantification by AE, the guidelines will recommend referral of bleeds >2ml for FC and instigation of follow‐up, even where the standard anti‐D dose exceeds 500 i.u.