Each of the purified rSEPs diluted in PBS to a concentration of 0.3 mg/ml was printed on epoxy slides (CapitalBio, Beijing, China) in 5 replicate spots as described previously [16 (link)]. Human IgG with serial dilutions (2.5, 5, 10 and 20 μg/ml) was used to fit the internal calibration curves or as positive controls. BSA in PBS or lysate of E. coli cells transformed with pET-32a plasmids at a concentration of 0.3 mg/ml was used as negative controls [16 (link)]. For quality control, the microarray slides were incubated with mouse anti-His tag IgG-Cy5 (SBA, Birmingham, AL) and the fluorescence intensity (FI) of each protein on the slides was scanned by GenePix Personal 4100A (Molecular Devices, Sunnyvale, CA) and analyzed by GenePix Pro 6.0 software (Molecular Devices, Sunnyvale, CA) [16 (link)]. Proteins with a signal-to-background ratio over 3.0 were used for further analysis [16 (link)].
Epoxy slides
Manufactured by CapitalBio
Epoxy slides are a type of laboratory equipment used for the preparation and analysis of samples. They provide a stable and durable surface for mounting and preserving samples for microscopic examination. Epoxy slides are designed to withstand various chemical and physical treatments, ensuring the integrity of the sample during the analysis process.
Automatically generated - may contain errors
2 protocols using epoxy slides
1
Microarray-based Immunogenicity Profiling
2
Antigen Microarray Assay for Antibody Selectivity
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Antibody selectivity was further investigated using an antigen microarray that contained 15 728 spots from 13 363 antigens representing 9882 unique Ensembl gene IDs. These antigens were protein fragments that were generated and verified within the Human Protein Atlas 9 . The array was produced in‐house as previously described 18 , 19 (Sjöberg et al., submitted for publication) and by printing antigens onto epoxy slides (CapitalBio) using a noncontact microarrayer (ArrayJet Marathon M025). The printed slide was dried, blocked, washed, and incubated with HPA029018 diluted to 0.2 μg/mL for 1 h RT. Secondary antibody anti‐rabbit IgG (Alexa Flour® 647) was applied to the array after washing and incubated for 1 h RT. The array was washed, spun dry, and scanned at 10 μm resolution using a microarray scanner (G2565BA, Agilent) and images were processed using software (GenePix 5.1, Molecular Devices) to collect the median intensities per spot at 633 nm.
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