Increased Retention of Cardiac Cells to a Glass Substrate through Streptavidin-Biotin Affinity.

Kara A Davis,Ahmed Abdel-Latif,Jonathan Satin,Brooke M Ahern,Brad J Berron,Andrea H Sebastian,Jensen Z Goh,Christine A Trinkle

doi:10.1021/acsomega.1c02003

Kara A Davis, Ahmed Abdel-Latif + Show 6 more

Open Access

https://doi.org/10.1021/acsomega.1c02003

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Abstract

In vitro analysis of primary isolated adult cardiomyocyte physiological processes often involves optical imaging of dye-loaded cells on a glass substrate. However, when exposed to rapid solution changes, primary cardiomyocytes often move to compromise quantitative measures. Improved immobilization of cells to glass would permit higher throughput assays. Here, we engineer the peripheral membrane of cardiomyocytes with biotin to anchor cardiomyocytes to borosilicate glass coverslips functionalized with streptavidin. We use a rat cardiac myoblast cell line to determine general relationships between processing conditions, ligand density on the cell and the glass substrate, cellular function, and cell retention under shear flow. Use of the streptavidin–biotin system allows for more than 80% retention of cardiac myoblasts under conventional rinsing procedures, while unmodified cells are largely rinsed away. The adhesion system enables the in-field retention of cardiac cells during rapid fluid changes using traditional pipetting or a modern microfluidic system at a flow rate of 160 mL/min. Under fluid flow, the surface-engineered primary adult cardiomyocytes are retained in the field of view of the microscope, while unmodified cells are rinsed away. Importantly, the engineered cardiomyocytes are functional following adhesion to the glass substrate, where contractions are readily observed. When applying this adhesion system to cardiomyocyte functional analysis, we measure calcium release transients by caffeine induction at an 80% success rate compared to 20% without surface engineering.

Highlights

In a critical effort to displace heart disease from its perennial position atop the leading causes of death in the developed world,[1] cardiac research has turned its focus to cellular function
The syncytial nature of the heart allows the use of isolated cardiomyocytes as surrogates for heart chamber function and for the interrogation of cellular and molecular mechanisms
Iwasaki and Ota showed that microarrays of functional, adhered cells could be achieved by patterning streptavidin on a silicon-based surface followed by incubation with biotinylated cells.[18]. We applied this simple concept by contacting biotinylated cardiac cells to streptavidinfunctionalized surfaces, and we observed a significant increase in cell immobilization despite high fluidic flow rates (Figure 1)

Summary

Introduction

In a critical effort to displace heart disease from its perennial position atop the leading causes of death in the developed world,[1] cardiac research has turned its focus to cellular function. The syncytial nature of the heart allows the use of isolated cardiomyocytes as surrogates for heart chamber function and for the interrogation of cellular and molecular mechanisms. L-type calcium channels in cardiomyocytes (CMs) are of great interest.[2−5] Multiple, interdependent processes regulate the sarcoplasmic reticulum Ca2+ load. Precise measurement of the Ca2+ load serves as a key integrative surrogate measure for heart function and prediction of disease. A standard bioassay of sarcoplasmic reticulum Ca2+ load requires CM immobilization onto an optically clear glass surface. CMs offer an added level of complexity compared to other primary cells due to their unique elongated shape and rigidity.[6]

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