Background: Immunogold cytochemistry is the method of choice for precise localization of antigens on a subcellular scale. The process of immunogold quantification in electron micrographs is laborious, especially for proteins with a dense distribution pattern.
New Methods: Here I present a MATLAB based toolbox that is optimized for a typical immunogold analysis workflow. It combines automatic detection of gold particles through a multi-threshold algorithm with manual segmentation of cell membranes and regions of interests.
Results: The automated particle detection algorithm was applied to a typical immunogold dataset of neural tissue, and was able to detect particles with a high degree of precision. Without manual correction, the algorithm detected 97% of all gold particles, with merely a 0.1% false-positive rate.
Comparisons With Existing Method(s): To my knowledge, this is the first free and publicly available software custom made for immunogold analyses. The proposed particle detection method compares favorably to previously published algorithms.
Conclusions: The software presented here will be valuable tool for researchers in neuroscience working with immunogold cytochemistry.
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http://dx.doi.org/10.1016/j.jneumeth.2017.05.018 | DOI Listing |
Nat Commun
September 2024
Department of Otolaryngology-Head & Neck Surgery, School of Medicine, 240 Pasteur Drive, Stanford University, Stanford, CA, 94305, USA.
Electron microscopy paired with immunogold labeling is the most precise tool for protein localization. However, these methods are either cumbersome, resulting in small sample numbers and restricted quantification, or limited to identifying protein epitopes external to the membrane. Here, we introduce SUB-immunogold-SEM, a scanning electron microscopy technique that detects intracellular protein epitopes proximal to the membrane.
View Article and Find Full Text PDFMicron
November 2024
Comparative Histolab Padova, Italy. Electronic address:
The process of keratinization and cornification in the developing beak has been studied through immunofluorescence and immunogold electron microscopy in chick and zebrafinch embryos. After the curved beak anlagen appears at the tip of the maxillar bone, 5-8 layers of embryonic epidermis are generated from the basal layer of the epidermis. These cells are weakly immunoabeled for IFKs (Intermediate Filament Keratins) and more intensely for scaffoldin, a protein of the EDC (Epidermal Differentiation Complex) involved in the soft keratinization of the embryonic epidermis.
View Article and Find Full Text PDFMethods Mol Biol
August 2024
Fischell Department of Bioengineering, University of Maryland, College Park, MD, USA.
Bacterial extracellular vesicles (BEVs) have emerged as mediators of transkingdom communication with numerous potential biotechnological applications. As such, investigation of BEV's protein composition holds promise to uncover new biological mechanisms, such as in microbiome-host communication or pathogen infection. Additionally, bioengineering of BEV protein composition can enhance their therapeutic potential.
View Article and Find Full Text PDFInt J Mol Sci
July 2024
Institute of Anatomy, Department of Neuroanatomy, Medical School, Saarland University, 66421 Homburg, Germany.
Synaptic ribbons are the eponymous specializations of continuously active ribbon synapses. They are primarily composed of the RIBEYE protein that consists of a unique amino-terminal A-domain and carboxy-terminal B-domain that is largely identical to the ubiquitously expressed transcriptional regulator protein CtBP2. Both RIBEYE A-domain and RIBEYE B-domain are essential for the assembly of the synaptic ribbon, as shown by previous analyses of RIBEYE knockout and knockin mice and related investigations.
View Article and Find Full Text PDFInt J Mol Sci
May 2024
Institute of Biology, Biotechnology and Environmental Protection, Faculty of Natural Sciences, University of Silesia in Katowice, 9 Bankowa St., 40-007 Katowice, Poland.
The genus (bladderworts) species are carnivorous plants that prey on invertebrates using traps with a high-speed suction mechanism. The outer trap surface is lined by dome-shaped glands responsible for secreting water in active traps. In terminal cells of these glands, the outer wall is differentiated into several layers, and even cell wall ingrowths are covered by new cell wall layers.
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