Human lactoferrin (aka lactotransferrin or growth-inhibiting protein 12) is an 82.4kDa 691 amino acid glycoprotein which carries 2 ferric ions per molecule and is the main iron binding protein in milk. Lactoferrin has multiple physiological functions including iron absorption regulation and antibacterial, antiviral, and antiparasitic activity. Degranulated neutrophils are the primary source of plasma lactoferrin which is increased during inflammation and bacterial infection. The sensitive quantitative measurement of human lactoferrin in human plasma, serum, milk, and cell culture samples is easily performed with this 96 well strip format ELISA kit. The normal human concentration of lactoferrin is 100 ng/ml in plasma and 5.3 mg/ml in milk. The assay measures human lactoferrin in the 0.1-100 ng/ml range. Samples giving human lactoferrin levels above 100 ng/ml should be diluted in blocking buffer before use. A 1:100-1:500 dilution for plasma or serum and and 1:200,000 dilution for breast milk is suggested for best results. Human lactoferrin will bind to the affinity purified capture antibody coated on the microtiter plate. After appropriate washing steps, biotinylated anti-human lactoferrin primary antibody binds to the captured protein. Excess primary antibody is washed away and bound antibody is reacted with peroxidase conjugated streptavidin. Following an additional washing step, TMB substrate is used for color development at 450nm. A standard calibration curve is prepared using dilutions of human lactoferrin and is measured along with the test samples. Color development is proportional to the concentration of total lactoferrin in the samples. All reagents and standards are provided in these ELISA kits.
