Antibody-modified reduced graphene oxide film for circulating tumor cell detection in early-stage prostate cancer patients

Abstract

In recent years, liquid biopsies, especially for detecting circulating tumor cells (CTCs), have received great attention for cancer diagnosis and treatment monitoring. For clinical diagnosis of prostate cancer (PCa), prostate specific antigen (PSA) has been widely used as a standard method for PCa screening. However, PSA diagnostic efficacy within early-stage PCa patients with a PSA level of 4-10 ng mL^-1 is always controversial. Therefore, the development of new methods to assist clinical PSA diagnosis is greatly desired. Herein, we report the fabrication of antibody-modified reduced graphene oxide films, which can be used to efficiently detect CTCs in PCa patients with PSA levels of 4-10 ng mL^-1. The antibody-modified reduced graphene oxide (rGO) films were fabricated by spray coating reduced graphene oxide solution onto a smooth glass slide and then modifying it with anti-epithelial cell adhesion molecules (anti-EpCAMs) and anti-prostate specific membrane antigen (anti-PSMA). The rGO films exhibited an excellent ability to capture CTCs from the blood of PCa patients with PSA levels of 4-10 ng mL^-1 and the efficiency could reach 60% (6/10). Our approach for highly efficient detection of CTCs in early-stage PCa patients may provide great potential in assisting clinical cancer diagnosis.

Fig. 1. Flow chart of rGO film fabrication and blood CTC detection. GO was reduced to rGO and rGO was printed onto glass slides as rGO film. The rGO films were functionalized using anti-EpCAM and anti-PSMA and were used for CTC detection in patients with PSA levels of 4–10 ng mL⁻¹.

Fig. 2. (A and E) ESEM images of the flat films with surfaces which do not have a microstructure and the PC3 cell, which displayed a spherical shape and does not have enough filopodia. (B–D) ESEM images of the rGO film surface which clearly show the existence of a petal-like wrinkled architecture. (F–H) The attachment relationship between a 22Rv1 cell, a PC3 cell and an mCRPC cell and the functional surface with the cell shape changed and the appearance of filopodia.

Fig. 3. (A) The capture efficiency for the PC3 cell line on rGO films and flat films. (B) The capture efficiency of PC3, 22Rv1, Daudi and Jurkat on anti-EpCAM- and anti-PSMA-modified films. (C) The capture efficiency of PC3 had a cell number-dependent linear relationship with R² = 0.9995.

Fig. 4. Immunofluorescence staining images, where CK⁺/CD45⁻ represents the CTC of a PCa patient and CK⁻/CD45⁺ represents the white blood cell of a PCa patient.