The invention discloses a method for detecting the drug sensitivity of traditional Chinese medicine monomer, which adopts the double dilution method of micro broth to determine the minimum inhibitory concentration of traditional Chinese medicine monomer acting on bacteria (including multi drug resistant bacteria), including the following steps: (1) dissolving the traditional Chinese medicine monomer in dimethyl sulfoxide, preparing the traditional Chinese medicine monomer solution with the concentration of 10N mg / ml; (2) taking 96 hole plate, adding 10 \u03bc l of dimethyl sulfoxide into each hole from the second row In the first column, 20 \u03bc l of Chinese medicine monomer solution was added; (3) multiple dilution was carried out successively; (4) the Chinese medicine monomer with different dose gradients was stored in the incubator until all dimethyl sulfoxide was volatilized; (5) the bacteria suspension with turbidity of 0.5 was added into 96 well plates, each well was 100 \u03bc L; the results were observed after culture at 37 \u2103 for 18h; (6) the results were determined: the minimum concentration without turbidity in the drug group was the minimum inhibitory concentration of Chinese medicine monomer Degree (MIC). The detection process of the invention is simple, simple and effective to test the drug sensitivity of the traditional Chinese medicine monomer, and can be used to guide the clinical treatment of bacterial infectious diseases with traditional Chinese medicine.
【技术实现步骤摘要】
一种中药单体药敏检测方法
本专利技术涉及药敏测试
,具体为一种中药单体药敏检测方法。
技术介绍
目前临床上对抗生素滥用的现象,对多重耐药菌的流行起到了一定的扩散作用。日益突出的多重耐药菌问题给临床上抗感染治疗带来了严峻的挑战。药敏实验为细菌感染性疾病的治疗,尤其是多重耐药菌感染的治疗提供了重要的参考价值。但是现今药敏实验全是西药抗菌药物的体外抑菌实验。近些年的研究报道发现有的中药成分对多种细菌,尤其是多重耐药菌具有抑制作用。但是临床仍缺乏商品化的临床简便可行而成本低廉的中药药敏实验器材。现中医治疗细菌感染性疾病仍是以传统辨证论治为主,缺乏精准治疗手段,临床疗效欠佳。本专利技术申请人在2019年温岭市科协“金桥工程”课题《黄芩苷对多重耐药菌的体外抑菌实验研究》的研究中,发现一种临床可行而成本低廉的中药单体抑菌实验方案,可为指导临床中药抗菌,尤其是对多重耐药菌的治疗提供重要的参考价值。
技术实现思路
为解决现有问题,本专利技术提供一种中药单体药敏检测方法。本专利技术通过以下技术方案实现。一种中药单体药敏检测方法,采用微量肉汤二倍稀释法测定中药单体作用于细菌(包含多重耐药菌)的最低抑菌浓度,包括以下步骤:(1)先将中药单体溶解于二甲基亚砜,配置成浓度为10Nmg/ml的中药单体溶液;(2)取96孔板,从第二列起每孔加入二甲基亚砜10μl,第一列加入步骤(1)配置的中药单体溶液20μl;(3)取在第一列细胞培养孔中取10μl加入到第二列细胞培养 ...
【技术保护点】
1.一种中药单体药敏检测方法,其特征在于:采用微量肉汤二倍稀释法测定中药单体作用于细菌(包含多重耐药菌)的最低抑菌浓度,包括以下步骤:/n(1)先将中药单体溶解于二甲基亚砜,配置成浓度为10N mg/ml的中药单体溶液;/n(2)取96孔板,从第二列起每孔加入二甲基亚砜10μl,第一列加入步骤(1)配置的中药单体溶液20μl;/n(3)取在第一列细胞培养孔中取10μl加入到第二列细胞培养孔中,依次进行倍比稀释;/n(4)倍比稀释后保存于温箱,直至二甲基亚砜全部挥发,细胞培养孔中剩余不同剂量梯度的中药单体;/n(5)将浊度为0.5的菌悬液加入96孔板,每孔100μl;最终中药单体浓度为N、N/2、N/4、N/8、N/16、N/32、N/64、N/128、N/256mg/mL;37℃培养18h后观察结果;/n(6)结果判定:药物组出现无浑浊的最小浓度为中药单体的最低抑菌浓度(MIC)。/n
【技术特征摘要】
1.一种中药单体药敏检测方法,其特征在于:采用微量肉汤二倍稀释法测定中药单体作用于细菌(包含多重耐药菌)的最低抑菌浓度,包括以下步骤:
(1)先将中药单体溶解于二甲基亚砜,配置成浓度为10Nmg/ml的中药单体溶液;
(2)取96孔板,从第二列起每孔加入二甲基亚砜10μl,第一列加入步骤(1)配置的中药单体溶液20μl;
(3)取在第一列细胞培养孔中取10μl加入到第二列细胞培养孔中,依次进行倍比稀释;
(4)倍比稀释后保存于温箱,直至二甲基亚砜全部挥发,细胞培养孔中剩余不同剂量梯度的中药单体;
(5)将浊度为0.5的菌悬液加入96孔板,每孔100μl;最终中药单体浓度为N、N/2、N/4、N/8、N/16、N/3...
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