Lectin from Bandeiraea simplicifolia (Griffonia simplicifolia)

Analysis Note

Agglutination activity is expressed in μg/mL and is determined from serial dilutions of a 1 mg/mL solution using phosphate buffered saline, pH 6.8, containing, for each lectin, calcium, magnesium, and manganese at different concentrations. This activity is the lowest concentration to agglutinate a 2% suspension of appropriate erythrocytes after 1 hr incubation at 25 °C.

Application

General Western Blot Protocol:
• Glycoprotein sample size: 500ng
• Lectin Concentration: 0.1ug/ml

1. Load samples at 500 ng of glycoprotein per lane
   • Run 4-20% Bis-Tris SDS page gel
   • Transfer gel to a PVDF membrane
   • Block membrane for 1 hr at RT with RIPA buffer (R0278 Sigma)
   • Incubate HRP lectin at 0.1ug/ml with RIPA buffer for 2 hours at RT
   • Wash membrane 5 x 5 minutes with 25ml RIPA buffer
   • Detect using chemiluminescent substrate (CPS1-120)

Biochem/physiol Actions

BS-I has a major affinity for terminal α-D-galactosyl residues with a secondary affinity for terminal N-acetyl-α-D-galactosaminyl residues.¹

Other Notes

BS-I is a tetrameric lectin consisting of two types of subunits designated A and B. There are five BS-I isolectins with different subunit composition: BSI-B₄, BSI-AB₃, BSI-A₂B₂, BSI-A₃B and BSI-A₄. BSI-B₄ is blood group B specific and has an exclusive affinity for terminal α-D-galactosyl residues, whereas BSI-A₄ has blood group A specificity and has a major affinity for terminal N-acetyl-α-D-galactosaminyl residues.

Preparation Note

Prepared from peroxidase type VI using a modification of the method of O′Sullivan, et al. Repurified by affinity chromatography after conjugation.

Unit Definition

One unit will form 1 mg purpurogallin in 20 sec from pyrogallol at pH 6.0 at 20 °C.

Physical form

Contains sodium citrate buffer salts and calcium chloride