Quantitation of D Sites on Selected 'Weak D' and 'Partial D' Red Cells

Abstract

Measurements have been made of the number of available sites on 10 examples of red cells in which the only abnormality appeared to be a quantitative reduction in the expression of D (weak D cells); these estimates were carried out using three monoclonal anti-D antibodies, Fog-1, Brad-3 and Los-2. The values varied with the monoclonal antibody that was used and fell within the range of 170-1,870 sites/cell. A further 3 examples of weak D cells which had brought about immunisation following transfusion were found to have between 390 and 1,470 sites per red cell. The implications of the D site density on the immunogenicity of weak D cells are discussed. The number of sites on red cells with structurally abnormal D (partial D cells) were also estimated, using the antibody Fog-1. Four of the 5 examples of cells of category IVa (probable phenotype R(0)r) were found to have a high expression of D (range 29,300- 41,300), but the available D sites of categories D^Va, D^VIa, and D^VII were considerably reduced (<500, <500 and 2,400-7,500 sites/cell, respectively). As a working hypothesis, it is suggested that there are two types of genetic abnormality leading to an abnormal expression of D. First, a defect in genomic DNA leading only to a quantitative reduction in the number of available D sites; this genomic lesion should be termed ‘weak D’. Secondly, genomic defects leading to amino acid sequence abnormalities and structural change in the D polypeptide; these lesions should be collectively known as ‘partial D’.

Quantitation of Rh D Antigen Sites on Weak D and D Variant Red Cells by Flow Cytometry

“…Not all reported D variants have been ex amined and a number of different techniques have been em ployed. The quantitative techniques employed include ag gregation [10][11][12], antibody coated rosette formation [13], radioisotopic labelling [14][15][16][17], ELISA [18] and flow cy tometry [19][20][21][22]. The number of D sites per cell (SPC) has been found to vary with different MAb-D for normal and weak D [17] and normal and D variant red cells [23], Cells belonging to the same D category also had different numbers of D SPC when quantified using the same anti-D [17,24].…”

Detection of weak D and DVI red cells in D‐negative mixtures by flow cytometry: implications for feto‐maternal haemorrhage quantification and D typing policies for newborns

“…In the Netherlands one incident was identified where a unit of weak D donor blood (1470 Rh D SPC), mistakenly typed as D negative, caused primary immunization in a female transfusion recipient with no previous history of pregnancy or transfusion (Gorick et al, 1993). For D VI blood there has been no reported incidences of immunization after transfusion in D-negative recipients.…”

“…If the fetus has a weak D or partial D VI phenotype much fewer sites may be present due to decreased Rh D antigen expression (Gorick et al, 1993;Jones et al, 1996). It has been estimated that the SPC in weak D vary between 100 and 1870 sites per cell, and in D VI between 300 and 2900, although a recently characterized D VI category (type III; Wagner et al, 1998) has been shown to be able to have near-normal Rh D antigen expression.…”

Principles and Applications of Red Blood Cell Flow Cytometry

“…Both direct or indirect immunofluorescence may be used. Flow cytometry has been applied to determine absolute antigen densities for ABO [7] and Rh [8][9][10]. The antigen density of weak D cells may be quantitized [8,10,11].…”

“…Flow cytometry has been applied to determine absolute antigen densities for ABO [7] and Rh [8][9][10]. The antigen density of weak D cells may be quantitized [8,10,11]. For other antigens, dosing effects and phenotype-related differences in antigen densities could be demonstrated [5,12].…”