Applied Biosystems™ PureQuant Monocyte Assay

Description

• The Applied Biosystems PureQuant assays are a family of assays that accurately identify and quantify specific immune cell types in a mixed population of cells
• These assays are TaqMan qPCR-based and measure genomic DNA methylation at specific sites that serve as a unique identifier
• The assays include standards and controls and each 96-well plate can accommodate up to seven test samples
• Results are provided as a percentage of target cell type out of total input cells using a free analysis tool
• The PureQuant Monocyte Assay utilizes the PARK2 gene as an epigenetic marker for monocytes
• Benefits include: Proven performance ―core technology has been used for identification and quantification of immune cell types for over a decade Easily standardized ―qPCR method enables consistent results across multiple users, instruments, and sites Facilitates regulatory filings ―regulatory support file provided upon request; established performance specifications following ICHQ2(R1) guidelines Scheduling flexibility ―assay can be performed using fresh or frozen cells/gDNA The assays include all reagents, standards, and controls required to run the assay
• Specific components include: Calibrator: corrects for variation in bisulfite conversion efficiency
• Plasmid-based calibrator harbors the genomic target regions in equimolar ratios and includes GAPDH
• Subjected to bisulfite conversion followed by copy number estimation via qPCR standard curve
• Allows estimation of assay-specific bisulfite efficiency factor: the ratio between TpG/GAPDH copy numbers measured from the same plasmid
• Reference: checks assay performance
• Consists of genomic DNA extracted from human blood of pooled healthy donors
• The reference values represent the expected range of the target cell type
• For example, CD8+ cells are generally more abundant than Treg or Th17 cells
• Standards: determine the unknown copy number
• Consist of six dilutions spanning 4-log linear dynamic range (30-31250 copies)
• Absolute copy numbers can reliably be determined using the slope and intercept of the respective standard curves
• This product is manufactured at a site with an ISO9001-certified quality management system.