Rapid detection of high-risk HPV16 and HPV18 based on microchip electrophoresis

Abstract

Researches on detection of human papillomavirus (HPV) high-risk samples were carried out by polymerase chain reaction (PCR) coupled with microchip electrophoresis (MCE). Herein, we introduced a simple, rapid, automated method for detecting high-risk samples HPV16 and HPV18. In this research, general primers were initially selected to obtain sufficient detectable yield by PCR to verify feasibility of MCM method for HPV detection, then type-specific primers were further used to evaluate the specificity of MCE method. The results indicated MCE method was capable of specifically detecting high-risk HPV16 and HPV18, and also enabled simultaneous detection of multiplex samples. This MCE method described here has been successfully applied to HPV detection and displayed excellent reliability demonstrating by sequencing results. The inherent capability of MCE facilitated HPV detection conducted in a small chip with automated, high throughput, massive parallelized analysis. We envision that MCE method will definitely pave a way for clinical diagnosis, and even on-site screening of cervical cancer.

Keywords: DNA analysis; Detection; Human papillomavirus; Microchip electrophoresis; Polymerase chain reaction.

Graphical abstract

Fig. 1

Schematic illustration of PCR combined with MCE for HPV detection. S, SW, B and BW indicate sample, sample waste, buffer and buffer waste, respectively.

Fig. 2

The microchip electrophoretograms of HPV16 and HPV18 detection. (A) The electrophoretogram of HPV16 detection by general primer PCR and optimized primer PCR. 1: DNA ladder; 2: general primer PGMY09/11 PCR product; 3: PGMY09-F/11-C PCR product. (B) The electrophoretogram of HPV18 detection by general primer PCR and optimized primer PCR. 1: DNA ladder; 2: general primer PGMY09/11 PCR product; 3: PGMY09-N/11-A PCR product.

Fig. 3

The electrophoretograms of HPV detection by type-specific primer PCR-MCE. 1: DNA ladder; 2: HPV16 DNA + HPV16 type-specific primer; 3: HPV18 DNA + HPV18 type-specific primer; 4: HPV16 DNA + HPV18 type-specific primer; 5: HPV18 DNA + HPV16 type-specific primer.

Fig. 4

The sensitivity of HPV detection. (A) The sensitivity of HPV16 detection by a serial 10 fold dilutions CaSki cell density (from 10⁶ to 10² CaSki cells/mL). 1, 2, 3, 4, 5 represent CaSki cell density, from 10⁶ to 10² cells/mL. (B) The sensitivity of HPV18 detection by a serial 10 fold dilutions Hela cell density (from 10⁶ to 10² Hela cells/mL). 1, 2, 3, 4, 5 represent Hela cell density, from 10⁶ to 10² cells/mL.

Fig. 5

Simultaneous detection of HPV16 and HPV18. (A) The simultaneous detection results of HPV16 and HPV18. (B) An enlargement of the gray part of figure A.