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Review Assays o Things to consider at first: Analyte: What kind of molecule is it?

t? MW, pH o (What kind of molecule it is) Small molecule do different style of test. Enzymes Concentrations: What concentrations are they in? Very High (ex. Plasma proteins) o Ex. Microwell EIA High (Antibodies) Low (Hormones) o Ex. Turbo o IFA Autoimmune Diseases Testing for antibodies Antimicrosomal, Antimicrobial, ENA Limited number of IFAs, generally have generic names such as antimicrosomal (not a specific protein, its a microsomal test) o If specifying antibody against specific protein in setting that its an autoimmune disease: We cant do that with an IFA because its not an individual protein, its a tissue on the slide. Has to be done in microwell add AB, let them bind, add conjugate, color change, etc. Cant do that on microscope slide (IFA) cant specify whats causing fluorescence o Hormones High sensitivity Steroid (SMALL) COMPETITIVE, ex. EMIT Thyroid or steroid hormones or small peptide hormones such as TRH Wont be able to get large antibodies to bind without inhibiting each other sterically. Hook one molecule onto enzyme and use EMIT (Competitive assay) instead of trying to get 2 ABs to bind 2 parts of small molecule. Large hormones (Peptide)- Sandwich o Enzyme Assays Generate peroxidase (color change from last step being peroxidase) OR use NADH reaction, 340nm for redox reaction o Electrodes Only 2 done by amperiometric: PO2 and glucose Everything else: Potentiometric, including free Calcium o Creatinine and Bilirubin Weird chemical assays o Iron and TIBC Fe H+ o Calcium and Magnesium and Phosphate All chemical assays React with something that turns color when bound o If it is a protein but its not an enzyme (no enzyme activity)? Treat the protein (analyte) like its an antigen Immunoassay almost always Is it one that is going to be in large quantities? Most often plasma proteins (large quantities) o OR Antibodies (large quantities) o OR peptide hormones (immunoassay but it will have to be highly sensitive turbo assay of some kind)