犬C-反应蛋白定量检测卡CRP

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【犬C-反应蛋白定量检测卡CRP】

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【公司名称】 广州健仑生物科技有限公司

【市 场 部】    杨永汉

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1.2.3.接种物制备 将用生长法或直接菌悬液法制备的浓度相当于0.5麦氏比浊标准的菌悬液，经MH肉汤1∶1000稀释后，向每孔中加100μl，密封后置35℃普通空气孵箱中，孵育16~20h判断结果。当试验嗜血杆菌属，链球菌属时，孵育时间为20~24h，试验葡萄球菌和肠球菌对苯唑西林和万古霉素的药敏试验时孵育时间必须满24h。此时，第1孔至第11孔药物浓度分别为128、64、32、16、8、4、2、1、0.5、0.25、0.125μg/ml。
1.2.4.结果判断 以在小孔内*抑制细菌生长的zui低药物浓度为MIC。当阳性对照孔（即不含抗生素）内细菌明显生长试验才有意义。当在微量肉汤稀释法出现单一的跳孔时，应记录抑制细菌生长的zui高药物浓度。如出现多处跳孔，则不应报告结果，需重复试验。通常对革兰阴性杆菌而言，微量肉汤稀释法测得的MIC与常量肉汤稀释法测得的结果相同或低一个稀释度（1孔或2倍）。
2.琼脂稀释法 琼脂稀释法是将不同剂量的抗菌药物，加入融化并冷至50℃左右的定量MH琼脂中，制成含不同递减浓度抗菌药物的平板，接种受试菌，孵育后观察细菌生长情况，以抑制细菌生长的琼脂平板所含zui低药物浓度为MIC。本法优点是可在一个平板上同时作多株菌MIC测定，结果可靠，易发现污染菌；缺点是制备含药琼脂平板费时费力。
2.1.培养基制备 使用MH琼脂，按商品说明书进行配制，pH7.2~7.4。淋病奈瑟菌使用GC琼脂基础加1%添加剂；其它链球菌使用含5%（V/V）绵羊血的MH琼脂（当试验磺胺药时，使用溶解的马血）。
2.2.含药琼脂平板制备 根据实验设计，将已倍比稀释的不同浓度的抗菌药物分别加入已加热溶解，并在45~50℃水浴中平衡的MH琼脂中，充分混匀倾倒灭菌平皿，琼脂厚度3~4mm。通常按1∶9比例配制药物琼脂平板，根据需要来选择药物浓度范围。配制好的含药琼脂平板应装入密封塑料袋中，置2~8℃冰箱可贮存5天。
2.3.接种物制备与接种 制备浓度相当于0.5麦氏标准比浊管的菌悬液，再1∶10稀释，以多点接种器吸取制备好菌液（约1~2μl）接种于琼脂平板表面，每点菌数约为104CFU，形成直径为5~8mm的菌斑。接种好后置35℃孵育16~20h（甲氧西林耐药葡萄球菌、万古霉素耐药肠球菌孵育时间应满24h），观察结果。奈瑟菌属、链球菌属细菌置5%二氧化碳、幽门螺杆菌置微需氧环境中孵育。
2.4.结果判断 将平板置于暗色、无反光物体表面上判断试验终点，以抑制细菌生长的zui低药物浓度为MIC。在含甲氧苄胺嘧啶或磺胺琼脂平板上可见轻微细菌生长，与生长对照比较抑制80%以上细菌生长的zui低药物浓度作为终点浓度。
如果出现有2个以上菌落生长于含药浓度高于终点水平的琼脂平板上，或低浓度药物琼脂平板上不长而高浓度药物琼脂平板上生长现象，则应检查培养物纯度或重复试验。

1.2.3. Inoculum preparation The growth suspension method or direct bacterial suspension concentration of the equivalent of 0.5 McNeil turbid standard bacterial suspension, diluted by 1: 1000 MH broth, to each well 100μl, Sealed at 35 ℃ normal air incubator, incubated 16 ~ 20h to determine the results. When testing Haemophilus, Streptococcus, incubation time is 20 ~ 24h, test staphylococcus and enterococci oxacillin and vancomycin susceptibility test incubation time must be full 24h. At this point, the first well to the 11th hole drug concentrations were 128,64,32,16,8,4,2,1,0.5,0.25,0.125μg / ml.
1.2.4. Results Judgment The minimum drug concentration to compley inhibit bacterial growth in the wells is MIC. When the positive control wells (that is, without antibiotics) significant bacterial growth test only makes sense. The highest drug concentration that inhibits bacterial growth should be recorded when there is a single perforated hole in the broth microdilution method. If there are multiple holes, you should not report the results, the need to repeat the test. For gram-negative bacilli, the MIC measured by the broth microdilution method is the same or a lower dilution (1 well or 2 times) as measured by the broth dilution method.
2. Agar dilution agar dilution method is to different doses of antibacterial drugs, added to melt and cooled to about 50 ° C quantitative MH agar, made with different decreasing concentrations of antibacterials of the plate, inoculated with test bacteria, bacteria were observed after incubation Growth conditions to inhibit the growth of bacteria in the agar plate containing the minimum concentration of MIC. The advantage of this method is that it can measure multiple strains of MIC at the same time on a flat plate, and the result is reliable and easy to find the contaminated bacteria. The disadvantage is that it takes time and effort to prepare the medicated agar plate.
2.1 Medium Preparation MH agar, formulated according to the product instructions, pH 7.2 ~ 7.4. Neisseria gonorrhoeae used GC agar base plus 1% additive; other streptococci used MH agar containing 5% (V / V) sheep blood (when sulfa drugs were tested, dissolved horse blood was used).
2.2. Drug-containing agar plate preparation According to the experimental design, different concentrations of antibacterial drugs that have been diluted to multiple dilutions were respectively added to MH agar which had been heated to dissolve and equilibrated in a water bath at 45-50 DEG C, Agar thickness of 3 ~ 4mm. The drug agar plate is usually formulated in a ratio of 1: 9, and the range of drug concentration is selected as needed. Prepared good containing agar plate should be sealed plastic bag, set 2 ~ 8 ℃ refrigerator can be stored for 5 days.
2.3 Inoculum preparation and inoculation Preparation of the equivalent of 0.5 Macrobrachium standard turbidimension bacterial suspension, and then 1:10 dilution, with a multi-point inoculum prepared bacteria (about 1 ~ 2μl) inoculated on the surface of the agar plate , The number of bacteria per point is about 104CFU, forming a diameter of 5 ~ 8mm plaque. After inoculation set 35 ℃ incubated 16 ~ 20h (methicillin-resistant staphylococcus, vancomycin-resistant enterococci incubation time should be full 24h), the observation results. Neisseria, Streptococcus bacteria set 5% carbon dioxide, Helicobacter pylori microaerophilic environment incubated.
2.4 determine the results of the plate placed on a dark, non-reflective surface to determine the end of the test to inhibit bacterial growth minimum drug concentration MIC. Slight bacterial growth was seen on trimethoprim or sulfanilamide agar plates, with the lowest drug concentration that inhibited the growth of more than 80% of the bacteria compared to the growth control as the endpoint concentration.
If more than 2 colonies grow on agar plates with drug concentrations above the end point or on long, high concentration drug agar plates on low concentration drug agar plates, check the culture for purity or repeat the test.