Single-cell protein profiling in microchambers with barcoded beads

Single-cell profiling provides insights into cellular behaviour that macroscale cell cultures and bulk measurements cannot reveal.In the context of personalized cancer treatment,the profiling of individual tumour cells may lead to higher success rates for therapies by rapidly selecting the most efficacious drugs.Currently,genomic analysis at the single-cell level is available through highly sensitive sequencing approaches.However,the identification and quantification of intracellular or secreted proteins or metabolites remains challenging.Here,we introduce a microfluidic method that facilitates capture,automated data acquisition and the multiplexed quantification of proteins from individual cells.The microfluidic platform comprises 1026 chambers with a volume of 152 pL each,in which single cells and barcoded beads are co-immobilized.We demonstrated multiplexed single-cell protein quantification with three different mammalian cell lines,including two model breast cancer cell lines.We established on-chip immunoassays for glyceraldehyde-3-phosphate dehydrogenase(GAPDH),galectin-3(Gal-3)and galectin-3 binding protein(Gal-3bp)with detection limits as low as 7.0×10^(4),2.3×10^(5)and 1.8×10^(3)molecules per cell,respectively.The three investigated cell types had high cytosolic levels of GAPDH and could be clearly differentiated by their expression levels of Gal-3 and Gal-3bp,which are important factors that contribute to cancer metastasis.Because it employed commercially available barcoded beads for this study,our platform could be easily used for the single-cell protein profiling of several hundred different targets.Moreover,this versatile method is applicable to the analysis of bacteria,yeast and mammalian cells and nanometre-sized lipid vesicles.

MyCTC chip:microfluidic-based drug screen with patient-derived tumour cells from liquid biopsies

Cancer patients with advanced disease are characterized by intrinsic challenges in predicting drug response patterns,often leading to ineffective treatment.Current clinical practice for treatment decision-making is commonly based on primary or secondary tumour biopsies,yet when disease progression accelerates,tissue biopsies are not performed on a regular basis.It is in this context that liquid biopsies may offer a unique window to uncover key vulnerabilities,providing valuable information about previously underappreciated treatment opportunities.Here,we present MyCTC chip,a novel microfluidic device enabling the isolation,culture and drug susceptibility testing of cancer cells derived from liquid biopsies.Cancer cell capture is achieved through a label-free,antigen-agnostic enrichment method,and it is followed by cultivation in dedicated conditions,allowing on-chip expansion of captured cells.Upon growth,cancer cells are then transferred to drug screen chambers located within the same device,where multiple compounds can be tested simultaneously.We demonstrate MyCTC chip performance by means of spike-in experiments with patientderived breast circulating tumour cells,enabling>95%capture rates,as well as prospective processing of blood from breast cancer patients and ascites fluid from patients with ovarian,tubal and endometrial cancer,where sensitivity to specific chemotherapeutic agents was identified.Together,we provide evidence that MyCTC chip may be used to identify personalized drug response patterns in patients with advanced metastatic disease and with limited treatment opportunities.