title: Aqueous two-phase system-mediated antibody micropatterning enables multiplexed immunostaining of cell monolayers and tissues.
abstract: Conventional immunostaining methods consume large quantities of expensive antibodies and are limited in terms of the number of antigens that can be detected from a single sample. In order to achieve multiplexed immunostaining, we micropatterned antibodies using aqueous two-phase systems (ATPS) formed from polyethylene glycol (PEG) and dextran. Multiple antigens can be detected on a single fixed sample by incorporating antibodies within dextran solutions, which are then patterned by micropipetting at specific sites on the sample in a solution of PEG. The antibodies are retained within the dextran phase due to biomolecular partitioning, allowing multiple protein markers to be visualized simultaneously by way of chromogenic, chemiluminescent, or immunofluorescent detection. This aqueous two-phase system-mediated antibody micropatterning approach allows antibody dilutions to be easily optimized, reduces the consumption of expensive primary antibodies and can prevent antibody cross-reactions, since the antibodies are retained at separate sites within the dextran microdroplets.
authors.affiliation: Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, USA.
authors.forename: John P
authors.initials: JP
authors.lastname: Frampton
authors.forename: Michael
authors.initials: M
authors.lastname: Tsuei
authors.forename: Joshua B
authors.initials: JB
authors.lastname: White
authors.forename: Abin T
authors.initials: AT
authors.lastname: Abraham
authors.forename: Shuichi
authors.initials: S
authors.lastname: Takayama
mesh_terms: D000818:Animals; D000906:Antibodies / Q000737:chemistry* / Q000378:metabolism; D002478:Cells, Cultured; D002645:Chickens; D003911:Dextrans / Q000737:chemistry; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods*; D061986:MCF-7 Cells; D008297:Male; D011092:Polyethylene Glycols / Q000737:chemistry; D017207:Rats, Sprague-Dawley
chemical_list: D000906:Antibodies; D003911:Dextrans; D011092:Polyethylene Glycols
keywords: Antibody micropatterning; Aqueous two-phase system; Immunohistochemistry; Multiplex immunostaining