DERC Mouse Phenotyping | The Histopathology Laboratory | Body Composition |
Multiplex Protein Assays (MPAs) | Whole Body Metabolic Assessment

Multiplex Protein Assays (MPAs)

A coherent analysis of circulating inflammatory proteins and metabolically relevant hormones has become a routine analytical tool for many DERC investigators. Measuring the concentration of multiple proteins in single, small volume samples (20-50 ul) is performed by the Mouse Phenotyping Core using a microsphere-based flow cytometry assay system developed by the Luminex Corp. The small volume required for accurate measurements and the multiplex nature of the assay permits DERC investigators to follow prospectively changes in circulating concentrations in key hormones, cytokines, chemokines, growth factors, angiogenic factors and inflammatory proteins in mice over time with minimal effort and without need for placement of indwelling catheters. The multiplex protein assay system has great flexibility and complements service provided by the analytical core, where assays are specifically tailored for individual proteins.

Location, Equipment and Staff:

The Mouse Phenotyping Core processes multiplex proteins assays using a Luminex 100 IS system (Austin, TX). This system is operated by a senior technician. The Luminex system is an open platform and therefore more than a dozen companies sell capture bead/secondary antibodies., such as Linco/Millipore, Invitrogen, R&D Systems and Qiagen. A single analysis run can measure 96 samples, and the maximum number of analytes per sample is currently 100, though most assays run typically measure ~7-10/sample. Many of the kits, such as Adipokine, Hormonal, and Metabolic, have pre-packed groups of capture beads/secondary antibodies that are relevant for diabetes researchers. For example, the Linco Standard Adipokine serum/plasma assay measures the concentration of: Il-6, leptin, PAI-1, MCP-1, Resistin, TNFa and insulin.

Procedure for using the Multiplex Protein Assay Laboratory:

DERC investigators with samples to analyze schedule time to meet with Natan Obstfeld using a web- based calendar. During the meeting, Mr. Obstfeld reviews the sample number request, the proteins to be assayed and the kit to be used. A time is then scheduled to perform the assay. The investigators purchase the capture bead/secondary antibody kit from an approved list of vendors and provide the samples (serum, plasma or conditioned medium) and assay kit to Mr. Obstfeld, who runs the assays. The samples are then analyzed, typically within three days of submission, and the data is provided to the investigator in spreadsheet form, following which an email to assess quality is sent to the investigator. Account information is collected and scheduling is arranged through the Columbia University DERC website. Users are charged based on samples analyzed (see price schedule below and on website).

Quality Control:

All users are mailed a short assessment questionnaire and are asked several simple questions. In addition, internal controls and diagnostics are run with each assay for each analyte. Maintenance service, including equipment quality assurance is provided by Luminex Corp (Austin, TX).