AFM-compatible microfluidic platform for affinity-based capture and nanomechanical characterization of circulating tumor cells

Abstract

Circulating tumor cells (CTCs) carried by the patient's bloodstream are known to lead to the metastatic spread of cancer. It is becoming increasingly clear that an understanding of the nanomechanical characteristics of CTCs, such as elasticity and adhesiveness, represents advancements in tracking and monitoring cancer progression and metastasis. In the present work, we describe a combined microfluidic-atomic force microscopy (AFM) platform that uses antibody-antigen capture to routinely isolate and nanomechanically characterize CTCs present in blood samples from prostate cancer patients. We introduce the reversible assembly of a microfluidic device and apply refined and robust chemistry to covalently bond antibodies onto its glass substrate with high density and the desired orientation. As a result, we show that the device can efficiently capture CTCs from patients with localized and metastatic prostate cancer through anti-EpCAM, anti-PSA, and anti-PSMA antibodies, and it is suitable for AFM measurements of captured intact CTCs. When nanomechanically characterized, CTCs originating from metastatic cancer demonstrate decreased elasticity and increased deformability compared to those originating from localized cancer. While the average adhesion of CTCs to the AFM tip surface remained the same in both the groups, there were fewer multiple adhesion events in metastatic CTCs than there were in their counterparts. The developed platform is simple, robust, and reliable and can be useful in the diagnosis and prognosis of prostate cancer as well as other forms of cancer.

Herringbone Microfluidic Probe for Multiplexed Affinity‐Capture of Prostate Circulating Tumor Cells

“…When spiked in blood, the capture purity of PC3 cells with reference to WBCs was ∼11%, a result comparable to previously reported values. [ 20,22 ] Moreover, the similarity in capture yields of both sets suggests that the HB‐MFP has the capacity to handle (with no clogging) viscous blood samples, and that the presence of WBCs, RBCs, and all other blood material has no significant effect on the multiplexed cell capture efficiency.…”

“…These values are within the range of previously reported sizes. [ 22,55,56 ]…”

“…Our previous efforts to apply reversible microfluidic chip proved successful in characterizing captured prostate CTCs using atomic force microscopy (AFM). [ 22 ] However, in the work, the transition from cell capture and identification to cell characterization was still susceptible to cell loss, which was associated with peeling off the PDMS. The HB‐MFP, on the contrary, is gently (in a controlled manner) pulled away from the capture substrate, permitting access of captured cells in a seamless way without loss or dislocation.…”

“…In our previous work, we showed that EpCAM, PSMA, and PSA antigens could be all effective for specific capture of CTCs. [ 22 ] Although successful, simultaneous processing of blood samples in three separate microfluidic chips limited the throughput for post‐capture identification of CTCs. The open‐space design of the HB‐MFP, on the other hand, paved the way to quickly hover blood samples over the activated antibody stripes and gave access to captured CTCs for their fluorescence staining without noticeable cell loss.…”

“…[ 21 ] However, existing challenges with these approaches mainly stem from associated high shear stresses within the narrow channels and restricted access to captured CTCs for downstream analysis. [ 22 ] In addition, common to these devices is the monoplexed cell capture, which limits the simultaneous use of multiple bio‐recognition ligands. [ 23 ] The Biocept platform [ 24 ] is an exception to this category, wherein a cocktail of multiple recognition molecules is deployed for cell capture.…”