OPFLP-08 Propagation and quantification of feline calicivirus using cell culture methods
|
ID： |
A3D5C776052B454BBD97ECCAE3DB2CB5 |
|
文件大小(MB)： |
0.09 |
|
页数： |
11 |
|
文件格式： |
|
|
日期： |
2012-3-3 |
|
购买： |
文本摘录（文本识别可能有误，但文件阅览显示及打印正常，pdf文件可进行文字搜索定位）：
Pu blished o n the Food D irec tora te’s (Health C anada) website at http://www.hc-sc.gc.ca/fn-an/res-rech/analy-meth/microbio/index-eng.php,Government of Canada Gouvernement du Canada,Laboratory Procedure OPFLP-08,MARCH 2010,HEALTH PRODUCTS AND FOOD BRANCH,OTTAWA,PROPAGATION AND QUANTIFICATION OF FELINE CALICIVIRUS USING CELL CULTURE METHODS,Kirsten Mattison1 Sabah Bidawid1, Carole Simard2, Louise Lessard2 Peter Mueller2, Yvon-Louis Trottier 3,1 Health Canada,Bureau of Microbial Hazards,251 Sir Frederick Bant ing Driveway,Ottawa,ON, Canada, K1A 0K9,2 Food Virology Centre of Expertise,St-Hyacinthe Laboratory,Canadian Food Inspection Agency ,3400 Casavant Boulevard West,J2S 8E3,3 Food Program,Regional Branch ,Health Canada,1001 St-Laurent Street West ,Longueuil , QC, Canada,J4K 1C7,Microbiological Methods Committee,Evaluation Division,Bureau of Microbial Hazards, Food Directorate,Postal Locator: 2204E,HPFB, Ottawa, Ontario, K1A 0K9,E-mail : micro.methods.committee@hc-sc.gc.ca,1. APPLICATION,This method describes how to perform all the steps involved in propagating, maintaining, detecting and,quantifying standardized stocks of the feline calicivirus (FCV) for use as a positive control in methods designed,to extract and detect RNA viruses of interest from food commodities.,2. DESCRIPTION,This method has been used successfully in our laboratories and can be easily transferred to allow all testing,laboratories to produce high quality stock s of FCV, providing appropriate facilities are available.,3. PRINCIPLE,Following procedures common to all viruses but tailored to fit the specific requirements of FCV, this procedure,describes how to grow and maintain stocks of Crandell-Reese Feline Kidney (CRFK) cells, and how to use,these cells to grow and quantify stocks of FCV. PCR detection and confirmation of FCV can be done using,OPFLP-06.,4. DEFINITION OF TERMS,4.1 See Appendix A of Volume 3.,4.2 Cytopathic Effect (CPE): The destruction of cells caused by viral infection. In CRFK cells infected with,FCV, CPE is characterized by cell rounding, lifting up from the m onolayer, and eventual cell lysis.,4.3 Plaque Forming Unit (PFU): A measurem ent of the concentration of infectious viruses in a sample.,Each plaque, or zone of clearing, in the monolayer is assumed to originate from one infectious virus,particle.,4.4 Multiplicity of Infection (MOI): The average number of virus particles present per cell. The MOI is,determined by dividing the number of infectious viral particles (previously determined as described in,section 7.8) added to a well or flask by the number of cells (determ ined as described in section 7.4).,OPFLP-08,March 2010,Pu blished o n the Food D irec tora te’s (Health C anada) website at http://www.hc-sc.gc.ca/fn-an/res-rech/analy-meth/microbio/index-eng.php,5. COLLECTION OF SAMPLES,5.1 This laboratory procedure is to develop standard reagents and should not necessitate sampling.,6. MATERIALS AND SPECIAL EQUIPMENT,Note: The Laboratory Supervisor must ensure that completion of the analysis, described in this method, must be,done in accordance with the International Standard referred to as "ISO/IEC 17025:2005 (or latest version).,General requirements for the com petence of testing and calibration laboratories".,SAFETY NOTE: USE OF CRYOTUBES IN THE LIQUID PHASE OF LIQUID NITROGEN MAY CAUSE,ENTRAPMENT OF LIQUEFIED NITROGEN INSIDE THE VIAL AND LEAD TO PRESSURE,BUILD-UP, RESULTING IN POSSIBLE EXPLOSION. WEAR GAUNTLETS AND EYE,PROTECTION AT ALL TIMES WHEN WORKING WITH LIQUID NITROGEN OR,MATERIALS STORED IN LIQUID NITROGEN.,Note: It is the responsibility of each laboratory to ensure that the incubators, block heaters or water baths are,maintained at the recommended temperatures. Where 37/C is recommended the waterbath must be at 37/C,+/- 1/C. For all other tem peratures it must be +/- 2/C.,6.1 Biological safety cabinet that provides a HEPA filtered work zone for protection of the reagents from,contamination.,6.2 Liquid nitrogen storage system or freezer capable of holding vials of cells (below -130/C).,6.3 Waterbaths capable of maintaining temperatures of 37/C and 45/C.,6.4 Incubator capable of maintaining a humidified environment (using a water tray) at 37/C with 5% CO2.,6.5 Freezers capable of maintaining -20/C and -70/C.,6.6 Inverted phase contrast m icroscope capable of visualizing tissue culture cells at 40X total magnification.,6.7 IEC DPR-6000 refrigerated centrifuge or equivalent.,6.8 IEC DPR-6000 rotors for 50 ml and 15 ml tubes or equivalent.,6.9 Hemacytometer with Neubauer rulings.,6.10 Microwave oven.,6.11 Autoclave.,6.12 Light box (for the reading of plaques).,6.13 Vor……
……