1. 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*,†,|

2. Introduction Integrated immunosorbent assay Microchip fabrication Experiment antigen and antibody –colloidal gold conjugated apparatus Result Discussion

3. Integrated immunosorbent assay *Conventional immunosorbent assay - need long time - many expensive reagent - troublesome procedure Integrated immunosorbent assay reduce assay time < 24 h simple operation

4. Microchip fabrication

5. Experiment Antigen and antibody-colloidal gold conjugate reaction : 50ul antiserum + 5ml of pH adjusted gold suspension (at room temp. for 30 min) blocking : 25ul 30% BSA in PB (at 4 C o overnight incubation) maximum absorbance at 525nm 1 week preserve without loss of activity Adsorption of s-IgA on polystyrene bead 50ul suspended bead + 0.5ml Ag solution (at room temp. for 1h and at 4 C o overnight incubation) blocking with casein solution (at room temp. for 1h ) washing PB

6. TLM (Thermal Lens Microscope) 진공상태나 Ar 가스 분위기 상태에서 재료를 고온으로 가열하여 thermal etching 및 gas etching 에 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 514.5nm probe beam : He-Ne laser 632nm

8. Result

9. Good point of this methods Specific interface ratio ( S / V ) 50ul micro plate ( 0.65mm diameter ) ==== 13cm -1 Microchannel (11bead 100um * 100um * 200um ) ==== 480cm -1 37 time larger than micro plate!! distance to reaction Microplate : 1.5mm Microchannel : 20um 5600 times shorter !! consequently reduce time to 10 min

10. Antigen adsorption -- Injection polystyrene bead -- s IgA sol (10ug/ml) injected -- incubation -- blocking casein -- AbCG sol injected for 10min almost all of reaction was finished in 20 min

11. Immunoassay in microchip Adsoption sample : 20 min blocking : 10 min Ag-Ab reaction : 10 min washing : 1min total less than 1 hour !!

12. Discussion Simple operation Remove troublesome procedure Reduce expensive reagent Association with automated system Application with DNA Computing