title: Rapid and sensitive electrochemiluminescence detection of rotavirus by magnetic primer based reverse transcription-polymerase chain reaction.
abstract: A novel method for detection of rotavirus has been developed by integrating magnetic primer based reverse transcription-polymerase chain reaction (RT-PCR) with electrochemiluminescence (ECL) detection. This is realized by accomplishing RT of rotavirus RNA in traditional way and performing PCR of the resulting cDNA fragment on the surface of magnetic particles (MPs). In order to implement PCR on MPs and achieve rapid ECL detection, forward and reverse primers are bounded to MPs and tris-(2,2'-bipyridyl) ruthenium (TBR), respectively. After RT-PCR amplification, the TBR labels are directly enriched onto the surface of MPs. Then the MPs-TBR complexes can be loaded on the electrode surface and analyzed by magnetic ECL platform without any post-modification or post-incubation process. So some laborious manual operations can be avoided to achieve rapid yet sensitive detection. In this study, rotavirus in fecal specimens was successfully detected within 1.5 h. Experimental results showed that the detection limit of the assay was 0.2 pg μL(-1) of rotavirus. The ECL intensity was linearly with the concentration from 0.2 pg μL(-1) to 400 pg μL(-1). What's more, the specificity of this method was confirmed by detecting other fecal specimens of patients with nonrotavirus-associated gastroenteritis. We anticipate that the proposed magnetic primer based RT-PCR with ECL detection strategy will find numerous applications in food safety field and clinical diagnosis.
authors: MOE Key Laboratory of Laser Life Science & Institute of Laser Life Science, College of Biophotonics, South China Normal University, Guangzhou 510631, China.
authors: Fangfang
authors: F
authors: Zhan
authors: Xiaoming
authors: X
authors: Zhou
authors: Da
authors: D
authors: Xing
mesh_terms: D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D006801:Humans; D008163:Luminescent Measurements / Q000191:economics / Q000379:methods*; D059346:Magnets / Q000737:chemistry; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000191:economics / Q000379:methods*; D048348:Reverse Transcription; D012401:Rotavirus / Q000235:genetics* / Q000302:isolation & purification*; D012400:Rotavirus Infections / Q000175:diagnosis* / Q000821:virology*; D012680:Sensitivity and Specificity; D013997:Time Factors
chemical_list: D017931:DNA Primers; D018076:DNA, Complementary; D012367:RNA, Viral