酶联免疫斑点法对抗结核治疗期肺结核患者多肽特异性IFN-γ的检测  

Detection of peptide-specific interferon-gamma with enzyme-linked immunospot assay in patients with pulmonary tuberculosis during the anti-tuberculosis treatment

作　　者：林淑娴[1] 杨芳芳[1] 彭毅[1] 李研[1] 方毅敏[2] 黎意芬[2] 赖小敏[1] 

机构地区：[1]中山大学中山医学院微生物教研室,热带病防治研究教育部重点实验室,海洋微生物功能分子广东省高校重点实验室,广东省重大传染病预防和控制技术中心,广州510080 [2]广州市胸科医院内科,510095

出　　处：《中国医药生物技术》2013年第2期93-99,共7页Chinese Medicinal Biotechnology

基　　金：十二五"传染病重大专项"分任务(2012ZX10004903-004-002);国家自然科学基金面上项目(81271779)

摘　　要：目的运用酶联免疫斑点法检测结核患者外周血IFN-γ,探讨结核患者结核多肽特异性IFN-γ分泌水平随抗结核治疗的变化。方法采用结核混合多肽E6+E7+C14、E6+E7作为刺激抗原,对331个处于治疗前或1、2、3、4、5和6个月治疗期的肺结核患者外周血标本进行多肽特异性IFN-γ酶联免疫斑点法(ELISPOT)检测。结果治疗前的肺结核患者其ELISPOT阳性检测率均高于痰涂片和痰培养阳性率(E6+E7+C14-ELISPOT92.5%、E6+E7-ELISPOT89.7%、痰涂片41.0%、痰培养59.0%),且差别均有统计学意义(P<0.010和P<0.001)。在抗结核治疗各个时期,肺结核患者IFN-γ-ELISPOT检测阳性率和斑点数(spotformingcells/106cells,SFP)随着治疗的进行持续下降,由治疗前高水平(E6+E7+C14-ELISPOT阳性率92.5%、SFP计数中值M=381;E6+E7-ELISPOT阳性率89.7%、SFP计数中值M=168)降至第6个月治疗结束时低水平(E6+E7+C14-ELISPOT43.6%、M=40;E6+E7-ELISPOT38.1%、M=36)。痰涂阳性患者的SFP计数持续处在高位,且治疗6个月痰涂阳性患者的SFP计数显著高于痰涂阴性患者。结论两种混合多肽IFN-γ-ELISPOT可提高结核患者的诊断率,同时检测抗结核治疗患者的IFN-γ分泌水平以监测抗结核治疗的效果。Objective To detect IFN-γ in the peripheral blood of pulmonary tuberculosis （PTB） patients by using IFN-γ enzyme-linked immunospot assay （ELISPOT） and investigate peptide-specific IFN-γ levels during anti-TB treatment. Methods Peptide-specific IFN-γ in peripheral blood （PLB） was detected by ELISPOT with Mycobacterium tuberculosis （MTB） peptide mixtures E6 ＋ E7 ＋ C14 and E6 ＋ E7 as stimulants. The PLB samples were taken from 331 PTB patients tracked longitudinally before treatment or 1, 2, 3, 4, 5, 6 months after treatment. Results Before treatment, the positive detection rates of two kinds of combined peptide mixture IFN-γ-ELISPOT were significantly higher than that of sputum smear and sputum culture （E6 ＋ E7 ＋ C14-ELISPOT 92.5%, E6 ＋ E7-ELISPOT 89.7%; sputum smear 41.0%, sputum culture 59.0%）. The positive detection rates and the spot forming cells per 106 cells （SFP） of IFN-γ-ELISPOT consistently decreased overall the treatment from higher level before treatment （by E6 ＋ E7 ＋ C14-ELISPOT, positive rate 92.5%, median SFP （M） = 381; by E6 ＋ E7-ELISPOT, positive rate 89.7%, M = 168） to lower level after six months treatment （by E6 ＋ E7 ＋ C14-ELISPOT 43.6%, M=40; by E6 ＋ E7-ELISPOT 38.1%, M = 36）. SFP values of patients with positive sputum smear were continued to remain high, and it was higher than that of patients with negative sputum smear aftet 6 months treatment. Conclusions The two combined peptide mixture IFN-y-ELISPOT assays increase the detection rates of PTB patients and monitor the efficiency of anti-TB treatment by detecting IFN-γ levels in patients with anti-TB treatment.

关 键 词：结核 肺 免疫酶技术 细胞 抗结核治疗 

分 类 号：R521[医药卫生—内科学]