Enzyme-Linked Lectinsorbent Assay Measures N -Acetyl-D-Glucosamine in Matrix of Biofilm Produced by Staphylococcus epidermidis

Abstract

An enzyme-linked lectinsorbent assay (ELLA) was developed for quantification of in situ biofilm produced by Staphylococcus epidermidis in polystyrene 96-well tissue culture plates with phosphatase-labeled wheat germ agglutinin (WGA) as a specific probe for the GlcNAcbeta-1,4n component of exocellular matrix material (EMM) that is responsible for intercellular adhesion and accumulation. The ELLA and the modified Christensen dye assay were used to test 13 laboratory strains of coagulase-negative staphylococci and 10 clinical isolates of S. epidermidis. Four biofilm-positive laboratory strains of S. epidermidis were positive by both tests, and six biofilm-negative strains were negative by both. One strain of S. hemolyticus was positive by the ELLA only. Two of the 10 clinical isolates of S. epidermidis were positive by both assays, two were negative by both, and the remaining were positive by the ELLA only. The ELLA was objective, reproducible, specific, sensitive, and useful for screening strains for their capacity to adhere to plastic, produce EMM, and form biofilm.

In situ cell and glycoconjugate distribution in river snow studied by confocal laser scanning microscopy

“…Fluorescent lectins have been suggested to distinguish between Gram-positive and Gram-negative bacteria (Sizemore et al 1990) and between different cell surface mutants of Rhodosporidium (Buck & Andrews 1999). Furthermore, lectins were used as a measure for biofilm matrix material in Staphylococcus biofilms (Thomas et al 1997).…”

Simple ELISA using Polyclonal Antibody and Lectin for Determination of Soluble Chitosan - Like Polysaccharide in Bacterial Culture

“…In this case, the presence of the competitive inhibitors can affect the binding of the lectin to the target molecule significantly. In fact, in the detection of GlcNAc in the biofilm matrix of Staphylococcus epidermidis by ELLA, GlcNAc inhibited the detection strongly [16]. Since the present method did not detected GlcNAc and the pentamer even at high concentrations (Figure 4), they might be washed out through the first capture reaction with the polyclonal antibody and the subsequent washing process.…”

“…Due to this specificity, GlcNAc and the oligomers work as strong competitive inhibitors for the binding of WGA to the target polysaccharides and glycoproteins [16,21,25,27]. The acetate medium used for the cultivation of the CP-producing strain contains yeast extract, which may provide GlcNAc and the oligomers from disrupted yeast cell walls.…”

“…WGA has been frequently used to detect the biofilm components (exopolysaccharides) of bacterial strains and the cell wall components (chitin/chitosan) in fungal and yeast cells [16,17,19,23,24], because WGA recognizes GlcNAc and its consecutive oligo units in polysaccharides [14,21,22]. Due to this specificity, GlcNAc and the oligomers work as strong competitive inhibitors for the binding of WGA to the target polysaccharides and glycoproteins [16,21,25,27].…”

“…https://scidoc.org/IJBMBS.php saccharides and glycoproteins [15][16][17][18][19][20]. Lectins can be obtained from many biosources such as bacteria, plants, and animals [14], and many lectins are commercially available.…”