title: Autoantigen Microarray Panel I
status: Public on May 21 2021
contact.city: POITIERS
contact.first: Mickaël
contact.last: MARTIN
contact.email: mickael-martin@outlook.com
contact.address: 2 rue de la Milétrie
contact.department: Internal Medicine
contact.country: France
contact.institute: Poitiers University Hospital
organism: Mus musculus
description: Array version: 98Ag_20171220 panel I
  Slide serial no.: UTPA20171220S34-35
manufacture_protocol: Serum samples were pretreated with DNAse-I and then diluted 1:50 in PBST buffer for autoantibody profiling. The autoantigen array bearing 94 autoantigens and 4 control proteins (diluted in PBS-0.1% Tween20 to 0.1 - 0.2 mg/mL) were printed in duplicates into Nitrocellulose film slides (FAST Slide, 16 pad, Whatman Inc). The diluted serum samples were incubated with the autoantigen arrays, and autoantibodies were detected with cy3-labeled anti-mouse IgG using a Genepix 4200A scanner (Molecular Device) with laser wavelength of 532 nm. The resulting images were analyzed using Genepix Pro 6.0 software (Molecular Devices).  The median of the signal intensity for each spot were calculated and subtracted the local background around the spot, and data obtained from duplicate spots were averaged. The background subtracted signal intensity of each antigen was normalized to the average intensity of the total mouse IgG, which was included on the array as an internal control. Finally, the net fluorescence intensity (NFI) for each antigen was calculated by subtracting a PBS control which was included for each experiment as negative control. Signal-to-noise ratio (SNR) was used as a quantitative measurement of the true signal above background noise. SNR values equal to or greater than 3 were considered significantly higher than background, and therefore true signals.