Hemagglutination assay

Hemagglutination is observed in the presence of staphylococci, vibrios, and other bacterial species, similar to the mechanism viruses use to cause agglutination of erythrocytes.

[3][4] The RBCs used in HA and HI assays are typically from chickens, turkeys, horses, guinea pigs, or humans depending on the selectivity of the targeted virus or bacterium and the associated surface receptors on the RBC.

A general procedure for HA is as follows, a serial dilution of virus is prepared across the rows in a U or V- bottom shaped 96-well microtiter plate.

The button appearance occurs because the RBCs are not held in the agglutinated lattice structure and settle into the low point of the U or V-bottom well.

The relative concentration, or titer, of the virus sample is based on the well with the last agglutinated appearance, immediately before a pellet is observed.

HI is closely related to the HA assay, but includes anti-viral antibodies as “inhibitors” to interfere with the virus-RBC interaction.

A standardized amount of virus or bacteria is added to each well, and the mixture is allowed to incubate at room temperature for 30 minutes.

HA and HI have the advantages that the assays are simple, use relatively inexpensive and available instruments and supplies, and provide results within a few hours.

The manual interpretation method introduces more opportunities for discrepancies in the assay because results can be subjective and the agreement between human readers is inconsistent.