方法部分补充练习_22832.docxVIP

MATERIALS AND METHODS (Role for Matrix Metalloproteinase 9 in Granuloma Formation during Pulmonary Mycobacterium tuberculosis Infection, from Cell, Vol. 148, Issue13, July. 2000)Mice. Specific-pathogen-free female, 6- to 8-week-old, CBA/J and C57BL/6 mice were purchased from Taconic (Germantown, NY), and wild-type FVB mice and MMP-9 null (MMP-9KO) breeder mice were purchased from Jackson Laboratory (Bar Harbor, ME). The MMP-9KO breeders were maintained as a colony at Colorado State University. All mice were maintained under barrier conditions with sterile mouse chow and water ad libitum. The specific-pathogen-free nature of the mouse colonies was demonstrated by testing sentinel animals, which were shown to be negative for 12 known mouse pathogens. All experimental procedures were approved by the Colorado State University Animal Care and Use Committee.小鼠 无特异病原体，6-8周大的 CBA/J 与 C57BL/6 雌性小鼠购自Taconic (Germantown, NY), 野生型FVB 小鼠与MMP-9 null (MMP-9KO) 育种小鼠breeder mice 购自Jackson实验室(Bar Harbor, ME)。MMP-9KO育种动物breeders 以群体形式as a colony 饲养于美国科罗拉多州大学。所有小鼠都隔栏under barrier conditions 喂养并随意喂无菌鼠食和水sterile mouse chow and water ad libitum。检测岗哨动物以证实小鼠群体的无特异病原体特征，结果表明12种已知的小鼠病原体均呈阴性。所有实验步骤均通过美国科罗拉多州大学动物保护使用委员会Colorado State University Animal Care and Use Committee同意.。 Infection with M. tuberculosis. Mycobacterium tuberculosis strain H37Rv, obtained from the Trudeau Mycobacterial Culture Collection, was passaged three times through pellicle and then stored frozen as a seed stock. Working stocks were made by passage through liquid culture of Proskauer-Beck medium containing 0.01% Tween 80, and aliquots of the organism in mid-log-phase growth were stored at 80°C until needed. Mice were infected using procedures described previously (4). Briefly, bacterial stocks were diluted in 5 ml of sterile distilled water to 2 x 106 CFU/ml and placed in a nebulizer attached to an airborne infection system (Glass-Col, Terre Haute, IN). Mice were exposed to 40 min of aerosol, during which approximatel