Isolation of chromosaponin I-specific antibody by affinity chromatography.
Chromosaponin I (CSI), a gamma-pyronyl-triterpenoid saponin isolated from pea and other leguminous plants, modulates several developmental processes of plant roots and activates the sugar taste receptor cells in blowflies. CSI is a unique saponin for its reducing power and biological activities in both plants and insects. In the present paper, we described the method of preparation for CSI-specific antibody using CSI-affinity and soyasaponin I-affinity columns. The antibody's-specific binding activity to CSI was confirmed by a bioassay using Arabidopsis roots and a ligand-molecule interaction analysis using BIAcore 3000. Because of the lability of CSI, the CSI-affinity column was made only by a moderate reaction condition in which CSI was coupled to EAH Sepharose 4B in the presence of 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC). The special control of the reaction temperature was essential to complete the coupling reaction; the reaction with EDC at 0 degrees C followed by a gradual increase in temperature.
Ahamed
Arifa
A
Graduate School of Science and Technology, Kobe University, Nada-ku, Kobe 657-8501, Japan.
Rahman
Abidur
A
Hayashi
Fumio
F
Ueji
Shinichi
S
Amakawa
Taisaku
T
Tsurumi
Seiji
S
D000906:Antibodies / Q000737:chemistry / Q000302:isolation & purification*; D017360:Arabidopsis / Q000378:metabolism; D001681:Biological Assay; D002845:Chromatography; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D005022:Ethyldimethylaminopropyl Carbodiimide / Q000494:pharmacology; D007700:Kinetics; D008024:Ligands; D008956:Models, Chemical; D011485:Protein Binding; D012503:Saponins / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification*; D012685:Sepharose / Q000494:pharmacology; D013696:Temperature; D013997:Time Factors
D000906:Antibodies; D008024:Ligands; D012503:Saponins; C075945:chromosaponin I; D012685:Sepharose; D005022:Ethyldimethylaminopropyl Carbodiimide