Human lactoferrin (LTF) is an 80kD glycoprotein which was first isolated from human milk. LTF is found in most body fluids and secretions, e.g., in the nose, genital tract, and tears.1 In the blood, LTF is secreted by neutrophils and its plasma concentration is positively related to the total pool of neutrophils and to the rate of neutrophil turnover.2 Because of its ability to strongly bind iron, LTF is considered to be bactericidal.3 In a number of cases of inflammation, LTF is released into the extracellular medium from secondary granules of neutrophilic leukocytes.4,5 Its extracellular concentration can therefore be used as an index of neutrophil activation, especially in blood samples containing anti-myeloperoxidase antibodies.
Principle of the Procedure:This method is an Enzyme-linked immunosorbent assay (ELISA) for lactoferrin. Lactoferrin is captured by a monoclonal antibody (MAb) that is coated on wells of a sectional microplate. A second LTF-MAb labeled with biotin is added to the well and binds with the captured LTF forming a “sandwich.” A solution of streptavidin-peroxidase is then added. Streptavidin has a high affinity for biotin and once bound, its HRP (HRP) label is available for color development by addition of the substrate, o-phenylenediamine (OPD). This color development at 450nm is proportional to the quantity of LTF in the sample.
Kit Format:6x16wells (96 assays)
Detection Limit:~2ng/ml
Kit Components:L1021-90A: Sample Dilution Buffer: PBS, BSA, Tween-20 and 0.01% thimerosal, 3 x 25mlL1021-90B: LTF Standard: Approximately 300ng of purified LTF with BSA in lyophilized form, 1x1vialL1021-90C: Wash Buffer (20x), Tris-HCl buffer containing NaCl, Tween-20, and 0.01% thimerosal. 1x100ml L1021-90D: LTF Antibody, Concentrated solution of monoclonal antibody to LTF in phosphate buffer containing NaCl, BSA, glycerol, and 0.01% thimerosal. 1x75ulL1021-90E: Streptavidin-HRP, Concentrated solution of streptavidin-coupled HRP in Tris-HCl buffer containing BSA and 0.01% thimerosal. 1x75ulL1021-90F: Dilution Buffer, PBS, BSA, Tween-20, and 0.01% thimerosal, 3x8mlL1021-90G: OPD Diluting Buffer, Citrate buffer containing H2O2 and 0.01% thimerosal. 1x20mlL1021-90H: Stop Solution, 1M H2SO4. 1x20mlL1021-90J: OPD, Four tablets
L1021-90K: Microtiter plate, 1x96 wells
Storage and Stability:Store *L1021-90B at 4°C; liquid at -20°C if not used within 3 days. Store other components at 4°C. Stable for at least 6 months. For maximum recovery of product, centrifuge the original vials after thawing and prior to removing the cap. L1021-90A and L1021-90F should be used within 5 days after opening of bottles.
