CAJ | 학술논문

目的建立白喉-破伤风-无细胞百日咳疫苗生产中百日咳毒素(pertussis toxin,PT)含量的测定方法.方法免疫家兔制备高效价抗PT血清,纯化抗PT多克隆抗体并进行酶标记,建立双抗体夹心ELISA法并对其进行验证.结果建立的方法在0～ 20 ng/ml PT测量区间呈现最佳线性,相关系数＞0.99,且重复性好.检测百日咳丝状血凝素和黏着素,结果均为阴性,说明该法特异性良好.试验内10次、不同试验间3次测定16.0、8.0、4.0、3.0 ng/ml PT,变异系数为2.8％～9.7％,回收率为95.7％～106.0％,精密度和准确度验证均符合常规质量控制要求,定量限度为3.0 ng/ml.结论该方法能有效检测百日咳杆菌大罐发酵过程中分泌的PT,为PT质量控制奠定了重要基础.
목적 건립백후-파상풍-무세포백일해역묘생산중백일해독소(pertussis toxin,PT)함량적측정방법.방법 면역가토제비고효개항PT혈청,순화항PT다극륭항체병진행매표기,건립쌍항체협심ELISA법병대기진행험증.결과 건립적방법재0～ 20 ng/ml PT측량구간정현최가선성,상관계수＞0.99,차중복성호.검측백일해사상혈응소화점착소,결과균위음성,설명해법특이성량호.시험내10차、불동시험간3차측정16.0、8.0、4.0、3.0 ng/ml PT,변이계수위2.8％～9.7％,회수솔위95.7％～106.0％,정밀도화준학도험증균부합상규질량공제요구,정량한도위3.0 ng/ml.결론 해방법능유효검측백일해간균대관발효과정중분비적PT,위PT질량공제전정료중요기출.
Objective To develop an ELISA method for quantitative determination of pertussis toxin (PT) during production of combined diphtheria,tetanus and acellular pertussis vaccine.Methods After chinchilla rabbits were immunized by PT,high titer anti-serum against PT was obtained and polyclonal antibody against PT was prepared.Then,a double antibody sandwich ELISA method was developed and verified.Results The best linearity ranged from 0 to 20 ng/ml of PT (correlation coefficient ＞ 0.99).No cross reactions with filamentous hemagglutinin and pertactin were observed by the developed ELISA.The variation coefficients of intra-and inter-assays were 2.8％-9.7％ and the recovery rates were 95.7％-106.0％ when 16.0,8.0,4.0 and 3.0 ng/ml of standard PTs were determined.The quantitative limit was identified as 3.0 ng/ml.Conclusion Due to the excellent specificity,precision,and accuracy,this method can be applied to detecting PT in fermentation broth quantitatively.