Direct protein microarray Fabrication using a hydrogel "stamper"

Langmuir

Volumn 14, Issue 15, 1998, Pages

  Martin, Brett D ^a   Gaber, Bruce P ^a   Patterson, Charles H ^a   Turner, David C ^a  

^a NAVAL RESEARCH LABORATORY   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

EID: 11544348184     PISSN: 07437463     EISSN: None     Source Type: Journal    
DOI: None     Document Type: Article

References (44)

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28. Polymerization was accomplished by addition of 0.05 wt % 2,2′-azobis[2-(2-imidazolin-2-ylpropane] initiator and 1.0 mol % methylene-bis-acrylamide cross-linker to an aqueous solution containing 15 wt % 6-acryloyl-β-O-methyl galactopyranoside. Amounts of initiator and cross-linker are relative to sugar monomer. The solution was introduced into the tubes, and the polymerization was permitted to proceed for 35 min under nitrogen at 55 °C, followed by cooling to 25 °C.
29. The overhead liquid level in the capillary was brought to 1 mm above the gel to maintain gel hydration. The loaded gel spontaneously extruded 20-50 μm.
30. 2. Coated plates stored longer than 1 week were not used.
31. The stamper approach, contact, and removal were observed under magnification using a CCD camera.
32. The stamped surface was rinsed vigorously with PBS for 30 s.
33. Blocking with BSA was undertaken via surface immersion in 0.1 wt % BSA in PBS for 30 min. The surface was not permitted to dry.
34. Typically, the stamped/blocked surfaces were immersed for 30 min in stirred (200 rpm) PBS at 25 °C containing 5 μg/mL fluorescein (FITC)-labeled antigen. After antigen exposure they were washed vigorously with PBS/0.5% Tween for 30 s and rinsed with PBS. Prior to visualization, coverslips were placed on the surface to maintain protein hydration.
35. Stamped antibodies and the corresponding labeled analogues were obtained from Sigma (St. Louis, MO).
36. "Background" refers to unstamped regions simply blocked with BSA.
37. For radiolabeling studies surfaces were prepared by stamping aminosilane-coated silicon wafer with a cubic stamper of dimensions 2 mm/side.
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