# An Optimized Langendorff-free Method for Isolation and Characterization of Primary Adult Cardiomyocytes

## TL;DR

This paper introduces a simplified method for isolating adult mouse heart cells without using the traditional Langendorff perfusion technique, making the process more accessible and easier for researchers.

## Contribution

The paper presents an optimized Langendorff-free method for isolating adult cardiomyocytes using simpler equipment and techniques.

## Key findings

- The Langendorff-free method eliminates the need for specialized equipment and cannulation.
- The method is suitable for researchers with limited resources or new to cardiomyocyte isolation.
- The protocol includes analysis of mitochondrial function in cardiomyocytes treated with LPS.

## Abstract

Isolation of adult mouse cardiomyocytes is an essential technique for advancing our understanding of cardiac physiology and pathology, and for developing therapeutic strategies to improve cardiac health. Traditionally, cardiomyocytes are isolated from adult mouse hearts using the Langendorff perfusion method in which the heart is excised, cannulated, and retrogradely perfused through the aorta. While this method is highly effective for isolating cardiomyocytes, it requires specialized equipment and technical expertise. To address the challenges of the Langendorff perfusion method, researchers have developed a Langendorff-free technique for isolating cardiomyocytes. This Langendorff-free technique involves anterograde perfusion through the coronary vasculature by clamping the aorta and intraventricular injection. This method simplifies the experimental setup by eliminating the need for specialized equipment and cannulation of the heart. Here, we introduce an updated Langendorff-free method for isolating adult mice cardiomyocytes that builds on the Langendorff-free protocols developed previously. In this method, the aorta is clamped in situ, and the heart is perfused using a peristaltic pump, water bath, and an injection needle. This simplicity makes cardiomyocyte isolation more accessible for researchers who are new to cardiomyocyte isolation or are working with limited resources. In this report, we provide a step-by-step description of our optimized protocol. In addition, we present example studies of analyzing mitochondrial structural and functional characteristics in isolated cardiomyocytes treated with and without the acute inflammatory stimuli lipopolysaccharide (LPS).