1970 nature 277 680 685 the lectin preparations were dialyzed extensively against phosphate buffered saline and filtered through 0 2 micron filters protein content was determined using micro bca kit and aliquots were kept at 4 u00b0 c the protein determination was frequently done to check protein loss if any cord blood was collected from local hospitals after obtaining informed consent of the mother the protocols described in these studies were approved by the institutional ethics committee in brief cord blood was collected in a sterile bottle containing heparin 40 iu of heparin ml of plain medium and processed soon thereafter by using standard protocols red blood cells rbcs were separated by hes 4 ml of blood 1 ml of hes and mononuclear cell mnc fraction was isolated by density gradient histopaque sigma using standard protocols mncs were suspended in isolation buffer and cd34 cells were isolated by immuno magnetic separation dynal as per the manufacturer 39 s instructions isolated cd34 cells were incubated with different concentrations 100 pg ml to 10 ng ml for all four lectins of lectins for various time periods 10 to 30 days this concentration range of lectins used was predetermined by carrying out toxicity tests on tf 1 cells using mtt or xtt assays the cd34 cells incubated for various time periods were subjected directly to cfu assay or seeded on irradiated m210b4 feeders for ltc ic assays as described.