Lectin Array 95

Identify and profile the glycans in their samples
Determine whether their biomarker of interest has glycan moieties
Find specific glycan binding ligands in biological samples

Blue ice

95

Label-basedSandwich-based

Semi-Quantitative

Dialysis Vials
Labeling Reagent
Labeling Buffer
Stop Solution
Lectin Array Glass Slide Assembly
Sample Diluent20
X Wash Buffer I20
X Wash Buffer IICy3 equivalent dye-conjugated Streptavidin
Slide Washer/Dryer
Adhesive device sealer
Floating Dialysis Rack
Manual

Dry the glass slide
Block array surfaces
Incubate samples (samples need to be biotinylated for the label-based approach)
For the the sandwich-based principle, incubate with a detection antibody cocktail
For the label-based principle, incubate the labeled-streptavidin.
Incubate with Cy3 Equivalent Dye-Streptavidin
Disassemble the glass slide
Scan with a gene microarray laser scanner
Perform densitometry and analysis

Quantitative analysis of lectin-glycoprotein interactions. Example: a concentration series of glycoproteins detected with the lectin array could reveal concentration dependent effects of lectin-glycan binding.Determine the profile of bacterial cell-surface glycans. Example: Cell lysate from bacteria can be Biotinylated and hybridized to the lectin array. Analysis of the binding pattern and correlation with the known carbohydrate-binding specificities of the lectins can determine the glycans on the cell membrane.

See Image/s
In this application, the RayBio Lectin Array 95 was used to detect specific glycosylations of purified Horseradish Peroxidase (HRP). Lectins BANLEC, BC2L-A, CALSEPA, GNA, HHA, NPA, PA-IIL, and PALa showed strong signals after incubation with 3.3 ug/mL Biotin-HRP followed by detection with streptavidin-fluorescence-dye (Figures A, B and C). The fluorescence signals from BANLEC, BC2L-A, CALSEPA, GNA, HHA, NPA, PA-IIL, and PALa were blocked in a concentration-dependent manner by HRP itself (Figures A and C), indicating that the signals were generated by lectin-HRP binding. These eight lectins are known to exhibit specific binding to mannose, which indicates that HRP contains mannose. After adding increasing amounts of mannose, the signals from BANLEC, BC2L-A, CALSEPA, GNA, HHA, NPA, PA-IIL, and PALa were reduced (Figures A and B).The reduction in signals from increasing concentrations of mannose confirms that HRP protein contains mannose in its glycocalyx. Additionally, the two lectins AAL and RS-FUC (fucose binding specificity) also showed strong interaction with HRP, which indicates the fucosylation of HRP. Overall, the results of the Lectin Array 95 were consistent with published literature regarding HRP glycosylation.