No abstract

An inhibitor of blood coagulation, a new protein with an apparent molecular weight of 34,000 and an isoelectric point of 4.9, was purified from human placental tissue by EDTA extraction. Five cDNA clones were isolated from the human placental lambda gt11 cDNA library using the mouse monoclonal antibody raised against the coagulation inhibitor as the probe. The longest insert consists of 1,566 nucleotides, and contains 960 nucleotides entirely encoding the 320 amino acids of the inhibitor, and a poly A tail. The deduced amino acid sequence was corroborated by chemical analyses of the protein. The entire amino acid sequence shows homology to those of lipocortin I, lipocortin II, and endonexin-related proteins. The cDNA for the inhibitor was expressed in Escherichia coli under the regulation of the trc promotor of the plasmid pKK233-2. The resulting recombinant protein manifested inhibitory activities against both blood coagulation and phospholipase A2 activity, as did the coagulation inhibitor isolated from human placenta.

Japanese cedar (Cryptomeria japonica) pollinosis has been reported to occur naturally in Japanese monkeys (Macaca fuscata) as well as humans. Using monoclonal antibodies (mAb) specific to Cry j 1, a major allergen in Japanese cedar pollen, we identified five independent epitopes (EP‐1 to EP‐5) on the molecule. The epitopes recognized by IgE antibodies in the sera of humans and monkeys with the pollinosis were analysed by an IgE enzyme‐linked immunosorbent assay inhibition method with these mAb. In human patients, the mAb to EP‐1 strongly blocked the binding of IgE antibodies in all patients’ sera to Cry j 1. The reaction patterns of IgE antibodies in monkeys, however, varied among the troops of monkeys. In some troops, the mAb to EP‐1 showed a blocking pattern similar to that for human patients. In other troops, mAb to EP‐4 and EP‐5 blocked binding of IgE. These results indicate that some, but not all, monkeys have antibody responses to the major allergen similar to those of humans.

We have devised a new method of counting pollen allergen particles, modified from the fluorescent immunoblotting technique of Schumacher et al. Airborne Japanese cedar pollen allergens collected on Burkard's sampling tape were transferred onto a nitrocellulose membrane. The membrane was then treated with antiallergen mouse monoclonal antibody conjugated with alkaline phosphatase. Pollen allergens were detected as purple spots on the nitrocellulose membrane after phosphate substrate staining was performed. Pollen allergen particles were visible under a stereoscopic microscope or to the naked eye and could thus be counted easily. This new counting method takes less time than previous methods and requires no special skill.

No abstract