Dilute
peptides, proteins, protein complexes, nucleic acids, or other antigens in PBS or H20
to 0.1 - 0.2 mg/mL (up to 30% DMF or DMSO can be used for peptides and
proteins that are insoluble in H20 or PBS). Diluted
antigens are transferred to 384-well "print plates" (7 μL/well).
Antibodies pre-labeled with Cy-3 and / or Cy-5 (diluted to 0.1 μg/μL)
are used as marker spots to orient the arrays post-scanning. Using
the robotic arrayer developed by Patrick
Brown (or equivalent) and CMP2 microarray
spotting pins (TeleChem International), antigen arrays are printed on
poly-L-lysine coated
microarray slides. For optimal results, increased pin sonication
between loads (5 x 7 second sonication cycles), between print plates (10
minutes), and at the end of each print (30 minutes) are recommended.
Arrays are stored at 4°in slide boxes sealed in plastic heat-sealable
pouches. Reactivity is generally best within 1 month of printing,
but many printed antigens retain excellent reactivity for months.
An
image of a robotic microarrayer assembled based on the design of Dr.
Patrick Brown. The microarrayer pictured resides in the laboratory of Dr.
Gary Schoolnik.
