title: Generation and application of ssDNA aptamers against glycolipid antigen ManLAM of Mycobacterium tuberculosis for TB diagnosis.
abstract: The development of effective Mycobacterial antigen diagnostic reagents remains a high priority. Mannose-capped lipoarabinomannan (ManLAM) is a lipoglycan serving as a major cell wall component. ManLAM is also an early released antigen in the blood circulation system during Mycobacteria tuberculosis (M.tb) infection and is a perfect target antigen for TB diagnosis. In this study, ssDNA aptamers "antibodies" against ManLAM of the predominant clinical epidemic M.tb Beijing genotype strains were generated by the Systematic Evolution of Ligands by Exponential Enrichment (SELEX) technique. The selected single aptamer T9 demonstrated the highest specificity and binding affinity, with an equilibrium dissociation constant (Kd) of 668 ± 159 nmol/L. We further detected ManLAM antigens in serum and sputum samples from active pulmonary tuberculosis (aPTB) patients, extrapulmonary TB (EPTB) patients and healthy donors by using a T9 based enzyme-linked oligonucleotide assay (ELONA). The results showed that the specificity and sensitivity were 95.31% and 83.00% (for 100 aPTB serum samples), 98.70% and 92.71% (for 96 aPTB sputum samples), and 94.44% and 88.71% (for 62 EPTB serum samples), respectively. A good correlation was observed between the T9 aptamer-based ELONA and the clinical T-SPOT.TB. Thus, T9 based ELONA has potentials for diagnosis of TB, including inactive TB, smear-negative TB, EPTB, and TB with immunodeficiency, and assist the diagnosis of LTBI albeit it could not distinguish LTBI and active TB.
authors: State Key Laboratory of Virology and Hubei Province Key Laboratory of Allergy and Immunology, Medical Research Institute, and Department of Immunology, Wuhan University School of Medicine, Donghu Road 165#, Wuhan 430071, Hubei Province, China; Department of Clinical Laboratory, the Second Hospital of Wuhu, Wuhu 241000, Anhui Province, China.
authors: Xiao-Lei
authors: XL
authors: Tang
authors: State Key Laboratory of Virology and Hubei Province Key Laboratory of Allergy and Immunology, Medical Research Institute, and Department of Immunology, Wuhan University School of Medicine, Donghu Road 165#, Wuhan 430071, Hubei Province, China; Wuhan Center for Clinical Laboratory, Puai Hospital, Tongji Medical College, Huazhong University of Science and Technology, China.
authors: Shi-Min
authors: SM
authors: Wu
authors: State Key Laboratory of Virology and Hubei Province Key Laboratory of Allergy and Immunology, Medical Research Institute, and Department of Immunology, Wuhan University School of Medicine, Donghu Road 165#, Wuhan 430071, Hubei Province, China.
authors: Yan
authors: Y
authors: Xie
authors: State Key Laboratory of Virology and Hubei Province Key Laboratory of Allergy and Immunology, Medical Research Institute, and Department of Immunology, Wuhan University School of Medicine, Donghu Road 165#, Wuhan 430071, Hubei Province, China.
authors: Neng
authors: N
authors: Song
authors: State Key Laboratory of Virology and Hubei Province Key Laboratory of Allergy and Immunology, Medical Research Institute, and Department of Immunology, Wuhan University School of Medicine, Donghu Road 165#, Wuhan 430071, Hubei Province, China.
authors: Qing
authors: Q
authors: Guan
authors: State Key Laboratory of Virology and Hubei Province Key Laboratory of Allergy and Immunology, Medical Research Institute, and Department of Immunology, Wuhan University School of Medicine, Donghu Road 165#, Wuhan 430071, Hubei Province, China.
authors: Chunhui
authors: C
authors: Yuan
authors: College of Chemistry and Molecular Sciences, Wuhan University, Wuhan 430072, Hubei Province, China.
authors: Xiang
authors: X
authors: Zhou
authors: State Key Laboratory of Virology and Hubei Province Key Laboratory of Allergy and Immunology, Medical Research Institute, and Department of Immunology, Wuhan University School of Medicine, Donghu Road 165#, Wuhan 430071, Hubei Province, China. Electronic address: zhangxiaolian@whu.edu.cn.
authors: Xiao-Lian
authors: XL
authors: Zhang
mesh_terms: D000293:Adolescent; D000328:Adult; D000942:Antigens, Bacterial / Q000032:analysis*; D052157:Aptamers, Nucleotide / Q000378:metabolism*; D002675:Child, Preschool; D005260:Female; D006801:Humans; D007223:Infant; D008070:Lipopolysaccharides / Q000032:analysis*; D008297:Male; D008875:Middle Aged; D009169:Mycobacterium tuberculosis / Q000302:isolation & purification*; D011446:Prospective Studies; D052156:SELEX Aptamer Technique; D012680:Sensitivity and Specificity; D044967:Serum / Q000737:chemistry; D013183:Sputum / Q000737:chemistry; D014376:Tuberculosis / Q000175:diagnosis*; D055815:Young Adult
chemical_list: D000942:Antigens, Bacterial; D052157:Aptamers, Nucleotide; D008070:Lipopolysaccharides; C050016:lipoarabinomannan
keywords: Aptamer; Diagnosis; Enzyme-linked oligonucleotide assay (ELONA); Mannosylated lipoarabinomannan (ManLAM); Mycobacteria tuberculosis (M.tb); Tuberculosis (TB)