Systematic comparison of surface coatings for protein microarrays.
To process large numbers of samples in parallel is one potential of protein microarrays for research and diagnostics. However, the application of protein arrays is currently hampered by the lack of comprehensive technological knowledge about the suitability of 2-D and 3-D slide surface coatings. We have performed a systematic study to analyze how both surface types perform in combination with different fluorescent dyes to generate significant and reproducible data. In total, we analyzed more than 100 slides containing 1152 spots each. Slides were probed against different monoclonal antibodies (mAbs) and recombinant fusion proteins. We found two surface coatings to be most suitable for protein and antibody (Ab) immobilization. These were further subjected to quantitative analyses by evaluating intraslide and slide-to-slide reproducibilities, and the linear range of target detection. In summary, we demonstrate that only suitable combinations of surface and fluorescent dyes allow the generation of highly reproducible data.
Guilleaume
Birgit
B
Department of Molecular Genome Analysis, German Cancer Research Center, 69120 Heidelberg, Germany. guilleaume@hd.rzpd.de
Buness
Andreas
A
Schmidt
Christian
C
Klimek
Fritz
F
Moldenhauer
Gerhard
G
Huber
Wolfgang
W
Arlt
Dorit
D
Korf
Ulrike
U
Wiemann
Stefan
S
Poustka
Annemarie
A
D000906:Antibodies / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry*; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate / Q000737:chemistry; D040081:Protein Array Analysis / Q000295:instrumentation / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013499:Surface Properties
D000906:Antibodies; D005456:Fluorescent Dyes; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate