Effects of lutropin (LH) receptor internalization and recycling on the apparent numbers of LH receptors in rat testis Leydig cells at different temperatures

The effect of different temperatures on the Scatchard analysis of binding studies with 125I-hCG and intact rat testis Leydig cells has been investigated. The results show that at temperatures greater than 4 degrees C an overestimation of the number of receptors/cell is likely to occur. A one-site analysis of the data gives values of 13,573, 61,924 and 3802 LH receptors/cell at 34 degrees C, 21 degrees C and 4 degrees C respectively. At 34 degrees C and 21 degrees C (but not at 4 degrees C) a two-site analysis of the data is possible, giving similar high affinity binding components at both temperatures (KD 1 X 10(-10) M) but with dissimilar low affinity binding components (34 degrees C, 2.17 X 10(-9) M and 21 degrees C, 6.3 X 10(-8) M). The calculated total number of LH receptors using a two-site analysis at 21 degrees C is equal to 114,766 receptors/cell, and at 34 degrees C it is 37,987 receptors/cell. It is proposed that the differences in the level of binding at different temperatures and the associated changes in the value of KD, reflect the temperature sensitivity of the endocytic pathway of the 125I-hCG/LH receptor complex within the rat testis Leydig cell. From previous studies it is known that at 34 degrees C and 21 degrees C, a large part of the cell-associated hormone is not receptor bound but rather it represents 'processed' hormone before it is released from the cell. At 4 degrees C the cell-associated 125I-hCG remains bound only to cell surface LH receptors and thus gives a more accurate measure of total receptor numbers/cell.     

荧光原位杂交技术分析人结肠菌群方法研究

建立荧光原位杂交技术分析人体内结肠菌群的方法。取受试者新鲜粪便 ,选用 5种特异性的 16SrRNA寡核苷酸探针 ,检测粪便样本收集后的保存时间、温度 ,离心条件及样本固定液存放时间对杂交计数结果的影响。结果建立最佳实验条件为 :粪便样本收集后应尽快在 4℃下保存 ,放置时间不要超过 12小时即作处理 ;样本的适宜离心条件为 70 0g 2分钟 ;样本用多聚甲醛固定后在 - 80℃下存放时间不要超过 5个月。该方法具有较好的稳定性 ,可以有效地检出个体之间结肠菌群的差异。