Uterine Environment and Breed: MATERIALS AND METHODS(2)

Hematological Measures—Fetal and Maternal

Blood samples were diluted in sterile heparinized saline, and total red blood cells (RBC) were counted using a he-mocytometer. To determine hematocrit percentage, small volumes of blood from Days 24 and 30 pooled samples and individual Day 40 and maternal samples were collected in heparinized microhematocrit capillary tubes (Scientific Products, McGaw, IL), centrifuged for 3 min in a microhematocrit centrifuge (Damon IED MB, Needham Heights, MA), and evaluated with a Lancer Spiracrit microhematocrit capillary tube reader (Brunswick Co., St. Louis, MO). Hemoglobin concentration was measured on pooled and individual blood samples in duplicate using a total hemoglobin kit (Sigma, St. Louis, MO). ventolin inhaler

Plots of expected vs. measured concentrations had slopes for maternal (b = 0.95) and fetal blood (b = 0.947) that did not differ from a value of one. Relative EPO concentrations in fetal and maternal plasma were measured with a heterologous RIA procedure (Diagnostics Systems Laboratories, Webster, TX) using a rabbit antiserum to human EPO. Duplicate 50-jjlI plasma samples were assayed in a final incubation volume of 100 |xl before addition of 0.5 ml of secondary antibody solution. EPO was measured in a single assay with an average intraassay coefficient of variation (CV) of 1.6%. The detection limit of the assay was 0.47 mU/tube. Serial dilutions of fetal and maternal plasma were parallel to the standard curve (data not shown). Accuracy of recovery for five known amounts (0.125-2.5 mU) of EPO added to maternal plasma was 93.6% (data not shown). Cross-reactivity with porcine EPO was not determined by the antibody supplier. However, the antibody does cross-react with rat (87%), sheep (45%), and mouse (39%) EPO. The antibody does not cross-react significantly with other hormones (e.g., human prolactin and insulin, rat FSH, hCG, or epidermal growth factor).


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