Abstract
Modern techniques for quantifying blood group antibody-antigen interactions are very limited, especially for weaker interactions which result from low antigen expression and/or partial expression of the antigen structure. Surface plasmon resonance (SPR) detection is often used to monitor and quantify bio-interactions. Previously, a regenerable, multi-fucntional platform for quantitative RBC phenotyping of normal antigen expression using SPR detection was reported. However, detection of weaker variants were not explored. Here, this sensitivity study used anti-human IgG antibodies immobilized to a gold sensor surface to two clinically important types of weaker D variants using SPR; weak D and partial D. Positive pre-sensitised cells bind to the anti-human IgG monolayer, and the response unit (RU) is reported (>100 RU). Unbound negative cells are directly eluted (<100 RU). Weak D cells were detected between a range of 180–580 RU, due to a lower expression of antigens. Partial D cells, category D VI, were also positively identified (352–1147 RU), similar to that of normal D antigens. The detection of two classes of weaker D variants was achieved for the first time using this fully regenerable SPR platform, opening up a new avenue to replace the current subjective and arbitrary methods for quantifying blood group antibody-antigen interactions.
Highlights
Mismatching incompatible blood types can lead to a haemolytic transfusion reaction, the severity of which can range from mild to fatal[1]
A CM5 sensor chip was functionalised with a monolayer of anti-human IgG prior to injection of red blood cells (RBCs) that have been sensitised with anti-D sera that is typically used For Further Manufacturing Use
The anti-D For Further Manufacturing Use (FFMU) selectively binds to the D antigen sites, allowing the Fc regions of the IgG antibodies covering the RBCs to be detected by the generic anti-human IgG immobilised onto the sensor chip surface
Summary
Mismatching incompatible blood types can lead to a haemolytic transfusion reaction, the severity of which can range from mild to fatal[1]. While the D antigen usually shows strong haemagglutination in the presence of the corresponding D antibody, there are weaker subgroup variants that do not react as strongly or as readily These interactions can be difficult to identify using traditional testing since blood group typing is dependent on a simple visual analysis, and can be overlooked or misinterpreted[2]. While much more difficult to identify because of their partial antigen or minute interactions, these weaker groups are as clinically significant, and can stimulate the formation of antibodies in the recipient which can still result in haemolytic transfusion reactions in subsequent transfusions. There are two methods available for quantitative analysis of RBC-IgG antibody interactions and antigen density which are not subjective to human interpretation; 1) flow cytometry, and 2) fluorescence microscopy. This was due to either the inability to fully desorb bound material or partial removal of the functionalised surface
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