Integration of an Immunosorbent Assay System: Analysis of Secretory Human Immunoglobulin A on Polystyrene Beads in a Microchip Kiichi Sato, † Manabu Tokeshi, † Tamao Odake, † Hiroko Kimura, ‡ Takeshi Ooi, § Masayuki Nakao, § and Takehiko Kitamori*, †, |
Introduction n n Integrated immunosorbent assay Microchip fabrication Experiment antigen and antibody –colloidal gold conjugated apparatus Result Discussion
Integrated immunosorbent assay *Conventional immunosorbent assay - need long time - many expensive reagent - troublesome procedure Integrated immunosorbent assay reduce assay time < 24 h simple operation
Microchip fabrication
Experiment n Antigen and antibody-colloidal gold conjugate reaction : 50 ul antiserum + 5 ml of p. H adjusted gold suspension (at room temp. for 30 min) blocking : 25 ul 30% BSA in PB (at 4 Co overnight incubation) maximum absorbance at 525 nm 1 week preserve without loss of activity n Adsorption of s-Ig. A on polystyrene bead 50 ul suspended bead + 0. 5 ml Ag solution (at room temp. for 1 h and at 4 Co overnight incubation) blocking with casein solution (at room temp. for 1 h ) washing PB
TLM (Thermal Lens Microscope) 진공상태나 Ar가스 분위기 상태에서 재료를 고온으로 가열하여 thermal etching 및 gas etching에 의하여 나타나는 미세조직의 변화를 관찰하고 그 온도에서 시료에 일어나는 여러 가지 변 화를 관찰. 금속, 세라믹 고분자 및 복합재료에서의 (1) 가열·냉각하는 동안의 상변태, (2)고온에서의 결정립의 성장, (3) 표면확산현상, (4) 고온 소성변형과 파 단, (5) 고온취성, (6) 금속 및 비금속의 소 결, (7)부식과정 등의 연구에 이용 excitation beam : Ar + laser 514. 5 nm probe beam : He-Ne laser 632 nm
Result
Good point of this methods Specific interface ratio ( S / V ) 50 ul micro plate ( 0. 65 mm diameter ) ==== 13 cm-1 Microchannel (11 bead 100 um * 200 um ) ==== 480 cm -1 37 time larger than micro plate!! distance to reaction Microplate : 1. 5 mm Microchannel : 20 um 5600 times shorter !! consequently reduce time to 10 min
Antigen adsorption -- Injection polystyrene bead -- s Ig. A sol (10 ug/ml) injected -- incubation -- blocking casein -- Ab. CG sol injected for 10 min almost all of reaction was finished in 20 min
Immunoassay in microchip Adsoption sample : 20 min blocking : 10 min Ag-Ab reaction : 10 min washing : 1 min total less than 1 hour !!
Discussion n n Simple operation Remove troublesome procedure Reduce expensive reagent Association with automated system Application with DNA Computing
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