Investigation of thiourea activated polyglutaraldehyde with bound Ag(I) or Pt(II) as an alternative to avidin for immobilizing biotin conjugates.
Pre-polymerized glutaraldehyde covalently linked to thiourea has been synthesized as a soluble polymer for immobilizing Ag(I) and Pt(II) and it has also been used for activating a polyacrylamide gel filtration media. The modified gel filtration media (Bio-Gel P-200) has a high capacity for Ag(I) (20 mumol/ml) and Pt(II) (8 mumol/ml) and has been shown to be stable and useful even in the presence of relatively high chloride (up to 1 M NaCl) and phosphate concentrations (0.25 M). The soluble polymer can have a Ag(I) capacity of between 2-11 mmol/g. Bio-Gel P-200 modified using glutaraldehyde/thiourea and in the Ag(I) and Pt(II) form selectively binds biotinylated BSA (b-BSA) over BSA. Using the Ag(I) form of the gel at pH 4.8 (0.05 M phosphate) only b-BSA binds and 30% can be eluted using 0.15 M NaCl, while no BSA binds to the column under these conditions. For the Pt(II) form of Bio-Gel P-200 at pH 4.8, none of the applied BSA binds to the modified resin while 40% of b-BSA does bind.
Department of Chemical, Bio & Materials Engineering, Arizona State University, Tempe 85287-6006, USA.
D H
DH
Kim
D R
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Miles
J W
JW
Knotts
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AA
García
D000418:Albumins / Q000737:chemistry; D001360:Avidin / Q000737:chemistry; D001710:Biotin / Q000737:chemistry*; D002021:Buffers; D002845:Chromatography / Q000379:methods; D003432:Cross-Linking Reagents / Q000737:chemistry; D004800:Enzymes, Immobilized / Q000737:chemistry*; D005782:Gels; D005976:Glutaral / Q000031:analogs & derivatives* / Q000737:chemistry; D010984:Platinum / Q000737:chemistry*; D011108:Polymers / Q000737:chemistry; D011485:Protein Binding; D012834:Silver / Q000737:chemistry*; D013890:Thiourea / Q000737:chemistry*
D000418:Albumins; D002021:Buffers; D003432:Cross-Linking Reagents; D004800:Enzymes, Immobilized; D005782:Gels; D011108:Polymers; D001360:Avidin; C022841:poly(glutaraldehyde); D012834:Silver; D010984:Platinum; D001710:Biotin; D013890:Thiourea; D005976:Glutaral