# Use of bovine serum albumin might impair immunofluorescence signal in thick tissue samples

## TL;DR

This study shows that using bovine serum albumin during tissue preparation can reduce fluorescence signal quality in thick, optically cleared tissues.

## Contribution

The study reveals that standard tissue blocking reagents, like bovine serum albumin, may impair fluorescence imaging in optically cleared thick tissues.

## Key findings

- Commonly used blocking reagents do not improve imaging conditions in optically cleared tissues.
- Bovine serum albumin can significantly degrade fluorescence signal quality in thick tissues like whole mouse brain hemispheres.

## Abstract

Significant progress in microscopic imaging techniques allowed transition from predominantly qualitative methods to a powerful tool for quantitative research, driven by improved instrumentation and computational power. Furthermore, previously limited to thin, laser-permeable tissue sections, imaging techniques have been revolutionized by the advent of tissue optical clearing. This innovation enables the visualization and quantitative analysis of entire organs and even whole bodies at cellular resolution. However, achieving high-quality imaging depends not only on the transparency of the tissue preparation but also on precise immunofluorescence labeling to ensure accurate signal detection and reliable study outcomes. In this study, we evaluated whether various reagents that are typically applied during the tissue blocking step prior to immunofluorescence staining affect the quality of the obtained image in thick and optically cleared samples. We demonstrate that the commonly employed tissue blocking step does not improve imaging conditions and even can substantially degrade fluorescence signal quality, particularly in large, optically cleared tissues such as whole mouse brain hemispheres.

The online version contains supplementary material available at 10.1038/s41598-025-06876-z.