Rapid typing of the human neutrophil antigen 1a by the particle gel agglutination assay.

The human neutrophil antigen 1a (HNA-1a) plays a major role in immune neutropenias and transfusion-associated lung injury. In this study, we describe a simple and rapid particle gel agglutination assay (PaGIA) for HNA-1a phenotyping. A commercially available monoclonal antibody (MoAb) to HNA-1a was biotinylated and coupled onto superparamagnetic streptavidin particles. Diluted anticoagulated whole blood samples from healthy blood donors (n = 147) were incubated with MoAb-coated particles, washed, transferred into an ID-gel card, and, subsequently, centrifuged. HNA-1a-positive samples resulted in a visible agglutination of the particles on top of the gel column and could be evaluated macroscopically. The results obtained by the new test were identical with those obtained by the polymerase chain reaction-sequence-specific priming technique that was performed in parallel. Seventy-four (50.3%) of the 147 samples were found to be HNA-1a positive. The HNA-1a PaGIA is both simple and safe and can be implemented in various laboratory settings.

Application of the particle gel agglutination system as a new check gel assay for PCR products.

In this study, we describe a simple and rapid agglutination test for the detection of PCR products prior to the application of specific hybridization by sequence-specific oligonucleotide typing. This test is based on the particle gel agglutination immunoassay, incorporating biotinylated primers and streptavidin particles. Visually detectable agglutination was only observed in samples which contained the specific amplification products. The results obtained by the new test were in accordance with those obtained by standard gel electrophoresis in all cases that have been tested to date.

Human platelet antigen genotyping by using sequence-specific primers and the particle gel agglutination assay.

BACKGROUND AND OBJECTIVES: Polymerase chain reaction using sequence-specific primers (PCR-SSP) is currently the most widely used technique for human platelet antigen (HPA) genotyping. Here, we describe a novel particle gel-agglutination technique for simplified visualization of the amplified products. MATERIALS AND METHODS: Biotinylated primers were used to amplify HPA-1, -2, -3, -4, -5, -6, and -15, and the PCR products were incubated with streptavidin particles. Fluorescein isothiocyanate (FITC)-labelled primers [amplifying a fragment of the human growth hormone (HGH) gene] and anti-FITC-coated particles were used as internal controls. Agglutination of the particles in or on top of the gel indicated specific amplification. A total of 100 samples from blood donors was tested by using this new technique and a standard PCR-SSP protocol. RESULTS: The use of biotinylated sequence-specific primers resulted in PCR products that agglutinated streptavidin particles, and the FITC-labelled HGH primers led to agglutination of anti-FITC-coated particles. Negative reactions were clearly distinguishable from positive reactions. The results of the particle gel agglutination method were in concordance with those of the electrophoretic visualization in all cases tested. CONCLUSIONS: The new particle agglutination method is reliable and easy to use.