Analysis of Knops blood group antigens on CR1 (CD35) by the MAIEA test and by immunoblotting

Abstract

Kna, McCa, Sla and Yka are red cell antigens of relatively high frequency, located on complement receptor 1 (CR1, CD35). Antibodies to these Knops system antigens are not uncommon. They are not haemolytic and do not reduce the survival of transfused incompatible red cells, but they are a nuisance in transfusion laboratories as they can cause an incompatible crossmatch and must be identified before they can be dismissed as clinically insignificant. Human red cell alloantibodies can be shown to be Knops system antibodies by the monoclonal-antibody-specific immobilization of erythrocyte antigens (MAIEA) test, using murine monoclonal anti-CR1. In addition to confirming that Kna, McCa, Sla and Yka are located on CR1, the MAIEA test was used to confirm that Csa is not on CR1. Red cells of the Helgeson phenotype, the null phenotype of the Knops system by conventional serological methods, have levels of Kna, McCa, Sla and Yka intermediate between those of alpha-chymotrypsin-treated cells (which lack Knops system antigens) and those of positive control cells. Level of expression of Knops system antigens is very variable and intensity of staining of immunoblots probed with monoclonal anti-CR1 correlated with strength of Knops system antigens, as determined by the MAIEA test. In individuals heterozygous for alleles producing different allotypes, separate bands representing each allotype on an immunoblot showed identical intensity of staining, suggesting that the quantity of CR1 on red cells is controlled, at least in part, by a locus independent of CR1. An analysis of CR1 on red cells of individuals who have made Knops system antibodies suggested that the Knops system antigens and the antibodies that detect them are complex and heterogeneous.

The monoclonal antibody‐specific immobilization of erythrocyte antigens assay (MAIEA) in the investigation of human red‐cell antigens and their associated membrane proteins

“…Antigens of relatively high frequency belonging to the Knops blood group system -Kn a , McC a , Sl a and Yk awere shown to be located on CR1 by immunoprecipitation (Moulds et al, 1991;Rao et al, 1991). This has been confirmed by MAIEA: absorbance values obtained with antibodies to each of these four antigens with either of two examples of anti-CR1 (BGU-3 and E11) were at least four times greater than those obtained with antigennegative cells (Petty et al, 1997). Blocking between 184 A. C. Petty et al ᭧ 1997 Blackwell Science Ltd, Transfusion Medicine, 7, 179-188 Fig.…”

“…This supports the observations of Moulds et al (1992), which illustrated that Helgeson cells had a low copy number of CR1 molecules per cell. Although a full validation of the technique was not performed, preliminary statistical data did suggest that MAIEA could be used quantitatively in the analysis of CR1 (Petty et al, 1997).…”

Molecular analysis of the York antigen of the Knops blood group system

“…One of the causes of a variable expression is a Hin dIII restriction length polymorphism, which is correlated to high or low expression of the CR1 gene in the Caucasian population 20 . For individuals with the Helgeson phenotype, which is often associated with difficult detection of CR1 expression, it has been shown that a very low copy number of protein is present on their RBCs 21 …”

“…20 For individuals with the Helgeson phenotype, which is often associated with difficult detection of CR1 expression, it has been shown that a very low copy number of protein is present on their RBCs. 21 Antibodies against the Knops antigens are high-titer, low-avidity (HTLA) antibodies. These antibodies react weakly and with variable strength in routine indirect antiglobulin tests but are still detectable in high dilutions of patient serum.…”

Blood Grouping Techniques

“…This method is particularly useful for identifying antibodies in the Knops system (Petty et al 1997). This method is particularly useful for identifying antibodies in the Knops system (Petty et al 1997).…”