An automated reading of semi-quantitative hemagglutination results in microplates: Micro-assay for plant lectins

Abstract

An automated, qualitative and semi-quantitative micro-assay for the detection of plant lectins also known as phyto-agglutinins (glycoproteins; have exciting applications in medicines) is described as an alternative to conventional assays. The method developed in this work is based on hemagglutination (HA) assay that can simultaneously detect the presence and concentration (titer) of lectins in as many as 96 different samples without the aid of an expert eye. We used rice (Oryza sativa L.) seeds for making clarified lectin extracts and Arabic gum as positive control in phosphate buffer saline; the method is applicable to all kinds and parts of the plants. Rabbit red blood cells were used in order to carry out the HA assay in a miniaturized experiment using U-welled microplates (MPs). 25 µL of plant extract is sufficient to carry out HA micro-assay at incubation temperature of 38°C for 20 min. The method was standardized with an expert eye and automated by using MP reader. Moreover, a standard curve for the direct interpretation of lectin concentration is also developed by conversion of absorbance values into titer. The method described will save time, material, labor, and simplify the rapid semi-quantification of plant lectins.

Keywords

• phyto-agglutinins
• hemagglutination assay
• hemagglutination titer
• microplate reader
• rice seed lectins