美国FDA细菌学分析手册第八版(BAM)产单核李斯特菌的血清学鉴定.docVIP

BAM: Serodiagnosis of Listeria monocytogenes January 2001 Bacteriological Analytical ManualChapter 11Serodiagnosis of Listeria monocytogenes Authors: Reginald W. Bennett and Robert E. Weaver Although serological confirmation is not necessary for regulatory identification of Listeria monocytogenes, it is useful for determining the prevalence of specific serotypes in epidemiological studies and for environmental recontamination tracking. Early attempts to confirm L. monocytogenes serologically were generally not successful because of cross-reactions with other organisms (1). Serology, therefore, should always follow cultural and biochemical identification (see Chapter 10). Serological assays of Listeria, reviewed by Gray and Killinger (4), include agglutination, precipitation, and complement fixation assays as well as automated fluorescent antibody techniques, using flow cytometry (3) and rapid method ELISA kits, which are based on monoclonal and polyclonal antibodies (5). However, these serological systems were developed to detect all Listeria spp. They have been used successfully to screen foods for the genus (see Chapter 10), but they do not differentiate L. monocytogenes from other Listeria species. This chapter presents procedures for the serological identification of L. monocytogenes by flagellar and somatic antigenic profiles (2), using agglutination as the serological tool. A. Equipment and materials Centrifuge Refrigerator (4-6°C) Steamer Inoculating needle Tubes, 6 x 50 mm Test tube rack (for 6 x 50 mm tubes) Dispenser, 25, 50, 100 μl, or comparable Water bath (48°C) Microscope slides B. Media1 and reagents2 EB motility medium (M48) Tryptose phosphate broth (TPB) (M168) Formaldehyde, 37% solution Formal saline, 0.5% Physiological saline, 0.85% (R63) McFarland No. 3 turbidimetric standard (R42) Somatic (O) antisera Flagellar (H) antisera C. Preparation of materials and media EB motility medium. Prepare as specified. Distribute 10 m1 amou