Harenberg J, Malsch R, Piazolo L, Huhle G, Heene D L
Department of Medicine, Faculty of Clinical Medicine Mannheim, University of Heidelberg, Germany.
Am J Vet Res. 1996 Jul;57(7):1016-20.
To address the problem of heparin binding to leukocytes of various animal species.
Leukocytes of the various species were incubated with fluorescent-labeled, low molecular mass heparin (LMMH). Fluorescence intensity on granulocytes, lymphocytes, and monocytes was analyzed by flow cytometry analysis.
Leukocytes were prepared from EDTA-anticoagulated blood of human subjects, rats, rabbits, dogs, pigs, and sheep 3 times.
The leukocyte populations were identified by their light scatter properties. In addition, phycoerythrin-labeled CD4, CD13, and CD14 antibodies were used to identify human lymphocytes, granulocytes, and monocytes; CD4, GR-1, and CD11b antibodies were used for mouse, and CD45, RP 3, and ED 9 antibodies were used for identification of rat leukocyte subpopulations.
Granulocytes, monocytes, and lymphocytes of all species bound LMMH in dose-dependent manner. Binding of LMMH-tyramine (tyr)-fluorescein-5-isothiocyanate (FITC) to granulocytes was higher in human subjects, rats, rabbits, dogs, and pigs, compared with binding to monocytes and lymphocytes. Mouse and sheep granulocytes did not bind more heparin than monocytes or lymphocytes. Binding of LMMH-tyr-FITC was reversible in the presence of unlabeled heparin of LMMH. More then 99% of human, rat, rabbit, dog, and sheep granulocyte populations were distinguished from monocytes and lymphocytes by means of their fluorescence intensity owing to LMMH-tyr-FITC. This separation was not obtained for mouse and pig granulocytes.
Evidence of specific heparin binding to granulocytes of many species indicates the relevance of fluorescent-labeled LMMH for biological investigations.
Binding of heparin and LMMH to granulocytes, lymphocytes, and monocytes may have a substantial role in atherosclerosis, inflammation, malignancy, and immunologic diseases.
解决肝素与不同动物物种白细胞结合的问题。
将不同物种的白细胞与荧光标记的低分子质量肝素（LMMH）一起孵育。通过流式细胞术分析粒细胞、淋巴细胞和单核细胞上的荧光强度。
从人类受试者、大鼠、兔子、狗、猪和绵羊的乙二胺四乙酸（EDTA）抗凝血液中制备白细胞3次。
通过光散射特性识别白细胞群体。此外，使用藻红蛋白标记的CD4、CD13和CD14抗体来识别人类淋巴细胞、粒细胞和单核细胞；使用CD4、GR-1和CD11b抗体来识别小鼠白细胞亚群，使用CD45、RP 3和ED 9抗体来识别大鼠白细胞亚群。
所有物种的粒细胞、单核细胞和淋巴细胞均以剂量依赖方式结合LMMH。与单核细胞和淋巴细胞相比，LMMH-酪胺（tyr）-异硫氰酸荧光素（FITC）与人类受试者、大鼠、兔子、狗和猪的粒细胞结合更高。小鼠和绵羊的粒细胞与单核细胞或淋巴细胞相比，结合的肝素并不更多。在未标记的LMMH肝素存在下，LMMH-tyr-FITC的结合是可逆的。由于LMMH-tyr-FITC，超过99%的人类、大鼠、兔子、狗和绵羊粒细胞群体可通过其荧光强度与单核细胞和淋巴细胞区分开来。小鼠和猪的粒细胞无法实现这种区分。
许多物种的粒细胞存在特异性肝素结合的证据表明荧光标记的LMMH在生物学研究中的相关性。
肝素和LMMH与粒细胞、淋巴细胞和单核细胞的结合可能在动脉粥样硬化、炎症、恶性肿瘤和免疫疾病中起重要作用。
