Construction of a novel peptide nucleic acid piezoelectric gene sensor microarray detection system.
A novel 2 x 5 clamped style piezoelectric gene sensor microarray has been successfully constructed. Every crystal unit of the fabricated gene sensor can oscillate independently without interfering with each other. The bis-peptide nucleic acid (bis-PNA) probe, which can combine with target DNA or RNA sequences more effectively and specifically than a DNA probe, was designed and immobilized on the surface of the gene sensor microarray to substitute the conventional DNA probe for direct detection of the hepatitis B virus (HBV) genomic DNA. Detection conditions were then explored and optimized. Results showed that PBS buffer of pH 6.8, an ion concentration of 20 mmol/liter, and a probe concentration of 1.5 micromol/liter were optimal for the detection system. Under such optimized experimental conditions, the specificity of bis-PNA was proved much higher than that of DNA probe. The relationship between quantity of target and decrease of frequency showed a typical saturation curve when concentrations of target HBV DNA varied from 10 pg/liter to 100 microg/liter, and 10 microg/liter was the watershed, with a statistic linear regression equation of I gC = -2.7455 + 0.0691 deltaF and the correlating coefficient of 0.9923. Fortunately, this is exactly the most ordinary variant range of the HBV virus concentration in clinical hepatitis samples. So, a good technical platform is successfully constructed and it will be applied to detect HBV quantitatively in clinical samples.
Chen
Ming
M
Department of Laboratory Diagnosis, Southwest Hospital, Third Military Medical University, Chong Qing 400038, P R. China.
Liu
Minghua
M
Yu
Lili
L
Cai
Guoru
G
Chen
Qinghai
Q
Wu
Rong
R
Wang
Feng
F
Zhang
Bo
B
Jiang
Tianlun
T
Fu
Welling
W
D001483:Base Sequence; D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods*; D002021:Buffers; D002460:Cell Line; D004279:DNA, Viral / Q000235:genetics / Q000302:isolation & purification; D004867:Equipment Design; D006515:Hepatitis B virus / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D006863:Hydrogen-Ion Concentration; D036103:Nanotechnology; D015341:Nucleic Acid Probes / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics*; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation / Q000379:methods*; D009994:Osmolar Concentration; D020135:Peptide Nucleic Acids / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics*
D002021:Buffers; D004279:DNA, Viral; D015341:Nucleic Acid Probes; D020135:Peptide Nucleic Acids