Session Speaker

Application Of Phytohaemagglutinin (Pha) Mitogenic Effect On Cytogentic And Cell Culture Procedure
Abolfazl-Movafagh
Iran

Introduction: Phytohaemagglutinin (PHA or phytohemagglutinin) is a lectin found in plants, especially beans. It is a lectin, obtained from the red kidney bean that binds to the membranes of T cells and stimulates metabolic activity, cell division, etc. PHA actually consists of two closely related proteins, called Leucoagglutinin (PHA-L) and PHA-E. The letters E and L point to the fact that these proteins agglutinate Erythrocytes and Leukocytes, respectively. It is found in the highest concentrations in uncooked red kidney beans and white kidney beans (also known as cannellini), and is also found in lower quantities in many other types of green beans and other common beans (Phaseolus vulgaris), as well as broad beans (Vicia faba) such as fava beans. It has a number of physiological effects and is used in medical research. In high doses it is a toxin. The object of this research work was to prepare Phytohaemagglutinin from kidney beans.

Material and Methods: The total preparation of Phytohaemagglutinin was done in a private laboratory. For testing the product of Phytohaemagglutinin, in each sample of normal individual,0.5-1.0 ml PB was obtained. For culture cells, 3-5 × 106 cells were cultured in 4 ml medium( RPMI 1640 ,Gibco-BRL Grand Island, NY,USA) supplemented with 15 per cent heat inactivated fetal bovine serum. 0.1 ml Phytohaemagglutinin was added and kept at 370 °C in an atmosphere containing 5% CO2. The processing of chromosome preparations from 72-h cultures was performed according to standard methods. Slides were examined with an Olympus model BH-2 light microscope. Eighty well-spread metaphases were analyzed for each subject. Karyotypes were described according to ISCN(1985).

Results: Cytogenetic studies were performed in 100 normal healthy individuals. Of them 95 were males and 41 females; The median age of these normal control at the time of cytogenetic examination was 34 years (range 14 to 72 years). Abnormal metaphases were obtained in 2(2%) in individual and the percentage of abnormal cells recorded within the range of 30%-37% in the present research work. The results of the remaining normal individuals (98 ) revealed normal G- banding karyotype. For each individual, eighty well-spread metaphases were analyzed according to ISCN(1985).

Discussion: Our result have indicated that preparation of fresh Phytohaemagglutinin at the time of cytogenetic and cell culture procedure reveals satisfactory score. The overall frequency of chromosomal analysis in our study was better when compared with treated commercial Phytohaemagglutinin as reported in the International prestigious journals. This difference in the results may be due to freshness of the Phytohaemagglutinin mitogenic protein prepared here and to the commercially aged product.