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Lysosomal Hydrolases

Lysosomal Hydrolases

Lysosomes are the site for the degradation of macromolecules and biological polymers released during membrane trafficking events such as secretory, endocytic, autophagic, and phagocytic pathways. The membrane-enclosed area of the lysosome, called the lumen, contains hydrolytic enzymes active in an acidic environment. These acid hydrolases are functional at a pH between 4.5 and 5 and are involved in cellular processes such as cell signaling, energy metabolism, restoration of the plasma membrane,...

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Related Experiment Video

Updated: Jan 15, 2026

Imaging Amyloid Tissues Stained with Luminescent Conjugated Oligothiophenes by Hyperspectral Confocal Microscopy and Fluorescence Lifetime Imaging

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Immunostaining Methods for Lysosomal Storage Disorder Research.

Ewa A Ziółkowska¹, Sophie H Wang¹, Hemanth R Nelvagal¹

¹Department of Pediatrics, Washington University in St. Louis, School of Medicine, St. Louis, MO, USA.

Methods in Molecular Biology (Clifton, N.J.)

|October 13, 2025

Summary

We optimized immunofluorescence staining protocols for analyzing lysosome structure and dysfunction in tissues. This method enhances sensitivity and allows simultaneous detection of multiple antigens for consistent, high-quality histological examination.

Keywords:

Histology Immunofluorescence Immunohistochemistry Primary antibody Secondary antibody Staining TrueBlack®

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Area of Science:

Histopathology
Cell Biology
Immunohistochemistry

Background:

Immunostaining visualizes proteins and antigens in tissues via antigen-antibody interactions.
Immunoperoxidase methods have been superseded by immunofluorescence for enhanced sensitivity and multiplexing.
Tissue autofluorescence and storage material can obscure cellular details in specific disease studies.

Purpose of the Study:

To present an optimized immunofluorescence staining protocol for histological analysis.
To enable high-quality, consistent quantitative results in tissue examinations.
To facilitate the study of lysosomal structure and dysfunction.

Main Methods:

Utilized immunofluorescence staining for enhanced antigen detection.
Developed a counterstaining method to mask tissue autofluorescence.
Applied the protocol to analyze lysosome structural organization.

Main Results:

Achieved superior sensitivity and simultaneous detection of multiple antigens.
Successfully masked interfering histofluorescence and autofluorescent storage material.
Enabled detailed analysis of lysosomal impact on cells and tissues.

Conclusions:

The optimized protocol serves as a standard for high-quality histological examinations.
Immunofluorescence offers significant advantages over immunoperoxidase methods.
This technique is crucial for understanding lysosomal disorders at a cellular and tissue level.