Tyramide signal amplification (TSA) is an enzyme-mediated method to enhance the immunohistochemical detection of protein, nucleic acid, or other molecules in situ.Here we describe immunofluorescent detection of a low-abundance cytokine receptor, interleukin-17 receptor B (IL17RB) in U2OS cells, using tyramide signal amplification. In addition, we present a tyramide signal amplification compatible double-color immunostaining protocol using primary antibodies from the same host species. Those applications allow detection of cellular proteins with enhanced sensitivity and add flexibility on primary antibody selection in double- or multicolor immunofluorescence staining assays.
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http://dx.doi.org/10.1007/978-1-0716-0247-8_7 | DOI Listing |
Anal Chem
January 2025
The School of Information Sciences and Technology, Northwest University, Xi'an 710127, P.R.China.
Digital fluorescence immunoassay (DFI) based on random dispersion magnetic beads (MBs) is one of the powerful methods for ultrasensitive determination of protein biomarkers. However, in the DFI, improving the limit of detection (LOD) is challenging since the ratio of signal-to-background and the speed of manual counting beads are low. Herein, we developed a deep-learning network (ATTBeadNet) by utilizing a new hybrid attention mechanism within a UNet3+ framework for accurately and fast counting the MBs and proposed a DFI using CdS quantum dots (QDs) with narrow peak and optical stability as reported at first time.
View Article and Find Full Text PDFTheranostics
January 2025
Department of Nephrology, China-Japan Friendship Hospital (Institute of Clinical Medical Sciences), Beijing, 100029, China.
The tertiary structure of normal podocytes prevents protein from leaking into the urine. However, observing the complexity of podocytes is challenging because of the scale differences in their three-dimensional structure and the close proximity between neighboring cells in space. In this study, we explored podocyte-secreted angiopoietin-like 4 (ANGPTL4) as a potential morphological marker via super-resolution microscopy (SRM).
View Article and Find Full Text PDFTalanta
December 2024
Marshall Laboratory of Biomedical Engineering, Shenzhen Key Laboratory for Nano-Biosensing Technology, Department of Biomedical Engineering, Medical School, Shenzhen University, Shenzhen, Guangdong, 518060, PR China. Electronic address:
Colorimetric enzyme-linked immunosorbent assays (CELISAs) have long been used for protein biomarker detection in diagnostics. Unfortunately, as confined by the monochromatic nature of detection signals and the limited catalytic activity of enzymes, CELISAs suffer from poor visual resolution and low sensitivity, hindering their effectiveness for early diagnostics in resource-limited settings. Herein, we report an ultrasensitive, high-visual-resolution CELISA (named PE-TSA-AuAg Cage-CELISA) that combines kinetically controlled growth of Ag in AuAg nanocages with poly-enzyme-boosted tyramide signal amplification (PE-TSA), enabling visual semiquantitative detection of protein biomarkers at attomolar levels with the naked eye.
View Article and Find Full Text PDFJ Lipid Res
December 2024
Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan. Electronic address:
Nuclear lipids play roles in regulatory processes, such as signaling, transcriptional regulation, and DNA repair. In this report, we demonstrate that nuclear lipids may contribute to Ki-67-regulated chromosome integrity during mitosis. In COS-7 cells, nuclear lipids are enriched at the perichromosomal layer and excluded from intrachromosomal regions during early mitosis but are then detected in intrachromosomal regions during late mitosis, as revealed by TT-ExM (expansion microscopy with trypsin digestion and tyramide signal amplification), an improved expansion microscopy technique that enables high-sensitivity and super-resolution imaging of proteins, lipids, and nuclear DNA.
View Article and Find Full Text PDFAnal Chem
January 2025
Zhejiang Cancer Hospital, Key Laboratory of Zhejiang Province for Aptamers and Theranostics, Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022, P. R. China.
As severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection posed a significant threat to public health and the global economy, in vitro diagnosis of the SARS-CoV-2 nucleocapsid protein proved to be an effective way for SARS-CoV-2 infection control in the past years. Tyramide signal amplification (TSA) has been extensively utilized in tissue imaging and pathological diagnosis owing to the powerful signal enhancement. However, the elevated "ALWAYS ON" fluorescence background limited the accuracy and sensitivity of the conventional TSA assay.
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