Summary. Background: Clinically significant age-related differences in the anticoagulation effect of heparin have previously been established in vitro as well as in different clinical settings in vivo. These differences were hypothesized to be due to the age-specific differences in binding of heparin to plasma proteins. Objectives: The aim of this project was to investigate global agerelated differences in heparin binding to plasma proteins. Patients/Methods: Heparin-binding proteins were identified by incubating heparin-coated magnetic beads with plasma samples from neonates, children and adults, and purifying the proteins that were bound to the beads in this reaction system. Results: These results provide the first preliminary evidence of age-related differences in the total number and concentration of proteins bound to heparin. The results also suggest, for the first time, that there are age-related differences of heparin binding to antithrombin and thrombin. Conclusions: The results of this study, although preliminary, support and contribute to the explanation of the mechanism of age-related differences in the effect of heparin observed previously in vitro and in vivo.

We describe a system in which proliferating human breast cancer cells are monitored by NMR spectroscopy for at least 6 days in basement membrane gel (BMG)1 threads. The cells are perfused under standard sterile cell culture conditions. 31P-NMR spectra obtained continuously for up to 64 h showed an increase in the signals owing to an increasing number of cells. Cell division in the BMG is easily observed by microscope or by the human eye as the gel opacifies. Spectra of cells in the BMG threads at 20% confluency show a more rapid signal increase than at 60% confluency. Cells grown in vivo in nude mice show a spectrum markedly similar to in vitro spectra in BMG threads, whereas the same cells in agarose threads lack peaks owing to Pi, glycerophosphocholine, and glycerophosphoethanolamine. With the high resolution obtained from this system we distinguished intracellular from extracellular Pi in vitro, and found that the intracellular pH is equal to that observed in the same cell line in vivo. This cell-BMG system is in effect a model tumor, but it is composed of a homogeneous cell population that can be observed indefinitely as the cells reproduce. The material needed is inexpensive, the technique is simple and efficient, and no adaptation of the spectrometer is required. This model will be useful for studying intracellular metabolism and the interaction of cells with the basement membrane.