| 1. |
- Pina, Ricardo, et al.
(författare)
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Ten Approaches That Improve Immunostaining : A Review of the Latest Advances for the Optimization of Immunofluorescence
- 2022
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Ingår i: International Journal of Molecular Sciences. - : MDPI. - 1661-6596 .- 1422-0067. ; 23:3
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Forskningsöversikt (refereegranskat)abstract
- Immunostaining has emerged as one of the most common and valuable techniques that allow the localization of proteins at a quantitative level within cells and tissues using antibodies coupled to enzymes, fluorochromes, or colloidal nanogold particles. The application of fluorochromes during immunolabeling is referred to as immunofluorescence, a method coupled to widefield or confocal microscopy and extensively applied in basic research and clinical diagnosis. Notwithstanding, there are still disadvantages associated with the application of this technique due to technical challenges in the process, such as sample fixation, permeabilization, antibody incubation times, and fluid exchange, etc. These disadvantages call for continuous updates and improvements to the protocols extensively described in the literature. This review contributes to protocol optimization, outlining 10 current methods for improving sample processing in different stages of immunofluorescence, including a section with further recommendations. Additionally, we have extended our own antibody signal enhancer method, which was reported to significantly increase antibody signals and is useful for cervical cancer detection, to improve the signals of fluorochrome-conjugated staining reagents in fibrous tissues. In summary, this review is a valuable tool for experienced researchers and beginners when planning or troubleshooting the immunofluorescence assay.
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| 2. |
- Téllez-Arreola, José Luis, et al.
(författare)
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Analysis of the MCTP Amino Acid Sequence Reveals the Conservation of Putative Calcium- and Lipid-Binding Pockets Within the C2 Domains In Silico
- 2022
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Ingår i: Journal of Molecular Evolution. - New York, NY, United States : Springer. - 0022-2844 .- 1432-1432. ; 90, s. 271-282
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Tidskriftsartikel (refereegranskat)abstract
- MCTPs (Multiple C2 Domains and Transmembrane region Proteins) are evolutionarily and structurally related to other C2 proteins, which are central to exocytosis and membrane trafficking; however, their specific function has been little studied. MCTPs are associated with endosomes and the endoplasmic reticulum and possess three C2 domains (C2A-C2C) and two transmembrane regions (TMRs) well conserved in different species. Here, we generated structural models of the MCTP C2 domains of C. elegans and analyzed their putative function by docking, which revealed that these domains possess Ca2+- and lipid-binding pockets, suggesting that MCTPs play a significant, calcium-dependent role in membrane physiology.
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