Immunocytometric quantitation of foeto-maternal haemorrhage with the Abbott Cell-Dyn CD4000 haematology analyser

Abstract

This study evaluated the extended use of a haematology analyser (Abbott Cell-Dyn CD4000) for the immunofluorescent enumeration of foeto-maternal haemorrhage (FMH) with fluorescein isothiocyanate-labelled monoclonal anti-RhD. Method performance was assessed with artificial FMH standards, and a series of 44 clinical samples. Within run precision was <15% (coefficient of variation, CV) for FMH volumes of 3 ml and above, 18.8% at an FMH volume of 2 ml and 31.7% at an FMH volume of 1 ml. Linearity analysis showed excellent agreement (observed FMH% = 0.98x expected FMH% + 0.02), and a close relationship (R(2) = 0.99) between observed and expected FMH percentages. The lower limit of quantification of the CD4000 (SRP-Ret) method with a maximum CV of 15% was 1.6 ml, and the limit of detection was <1 ml. Parallel Kleihauer-Betke test (KBT) assessments of FMH standards showed an overall trend for higher KBT values (observed = 1.25x expected - 0.38). At an FMH level of 4 ml, KBT observer estimates ranged from 0.57 to 11.94 ml with a mean inter-observer CV of 63%. For 44 clinical samples, there was decision point agreement between KBT and SRP-Ret results for 42 samples with an FMH of <2 ml. Analysis in the low FMH range (<1 ml) showed that small volume foetal leaks could be detected with the SRP-Ret method in most of 23 samples with negative KBT results. CD4000 SRP-Ret method performance for FMH determination was similar to that reported for flow cytometry.

Detection of fetomaternal hemorrhage

“…With around‐the‐clock availability and a short 30‐minute turn‐around time, confirmatory FMH testing can be integrated into routine hematology testing menu. Equivalent results to flow cytometry have been shown for both the anti‐HbF and anti‐D antigen methods using one model [ 53]. With the trend of integrating extended applications into routine hematology analyzers, the automated reticulocyte count being a notable example, additional platform options and broader clinical use is predicted to follow.…”

“…They reported excellent agreement and correlation ( R 2 =0.99) between observed and expected fetal cell percentages. Additionally, the predicted lower limit for the quantitation of FMH was 1.6 mL with a maximum CV of 15%, which is well‐below the 15 mL threshold for additional RhIg dosing [ 53]. The application of a hematology analyzer for the quantitation of FMH is especially appealing, as it is fully automated, precise, and has a wide range of linearity.…”

Immunophenotypic analysis of cerebrospinal fluid cell populations with the Cell‐Dyn Sapphire haematology analyser: method feasibility and preliminary observations

“…Comparability and equivalence of Cell‐Dyn and flow cytometry fluorescent measurements are evidenced by many recent publications detailing myeloid cell, lymphocyte, platelet and red blood cell immunofluorescent applications (Arroyo et al. , 2001; Little et al. , 2005; Molero et al.…”

“…Fluorescence is an integral part of the CD-Sapphire and has the ability to replicate many of the basic analyses currently provided by conventional flow cytometers. Comparability and equivalence of Cell-Dyn and flow cytometry fluorescent measurements are evidenced by many recent publications detailing myeloid cell, lymphocyte, platelet and red blood cell immunofluorescent applications (Arroyo et al, 2001;Little et al, 2005;Molero et al, 2005;Johannessen et al, 2006;Yamane et al, 2006;Van der Meer et al, 2006;Molero et al, 2007).…”

Quality counts: new parameters in blood cell counting

“…The instrument can be used to measure foeto‐maternal haemorrhage, a rapid and cost effective alternative to the traditional Kleihauer–Betke technique which has been traditionally the only option available to most laboratories. The method requires a preincubation of prepared supernatant with FITC conjugated BRAD – three monoclonal anti‐D for 30 min and it is then processed on the Cell‐Dyn Sapphire (Little et al. , 2005).…”