Localization of Epitopes for Monoclonal Antibodies Directed against the Adult Rat Skeletal Muscle Sodium Channel (rSkM1) Using Polymerase Chain Reaction, Fusion Proteins, and Western Blotting

1995 ◽  
Vol 226 (1) ◽  
pp. 188-191 ◽  
Author(s):  
W.J. Sun ◽  
S.A. Cohen ◽  
R.L. Barchi
Keyword(s):  
Skeletal Muscle ◽  
Polymerase Chain Reaction ◽  
Monoclonal Antibodies ◽  
Sodium Channel ◽  
Western Blotting ◽  
Fusion Proteins ◽  
Rat Skeletal Muscle ◽  
Chain Reaction ◽  
Adult Rat ◽  
Polymerase Chain

Tissue specificity and alternative splicing of the Na+/Ca2+ exchanger isoforms NCX1, NCX2, and NCX3 in rat

1997 ◽  
Vol 272 (4) ◽  
pp. C1250-C1261 ◽  
Author(s):  
B. D. Quednau ◽  
D. A. Nicoll ◽  
K. D. Philipson
Keyword(s):  
Skeletal Muscle ◽  
Polymerase Chain Reaction ◽  
Alternative Splicing ◽  
Reverse Transcriptase ◽  
Splice Variants ◽  
Variable Region ◽  
Intracellular Loop ◽  
Chain Reaction ◽  
Splicing Isoforms ◽  
Polymerase Chain

The gene coding for the Na+/Ca2+ exchanger NCX1 is characterized by a cluster of six exons (A, B, C, D, E, and F) coding for a variable region in the COOH terminus of the large intracellular loop of the protein. Alternative splicing of these exons generates multiple tissue-specific variants of NCX1. Using reverse transcriptase-polymerase chain reaction, we analyzed eight previously described and four new splicing isoforms of NCX1 in a wide variety of tissues and cells. Exons A and B are mutually exclusive, as shown in earlier studies, and splicing isoforms containing exon A are preferentially expressed in heart, brain, and skeletal muscle, whereas splicing variants with exon B are found in all rat tissues except heart. The second and third isoforms of the Na+/Ca2+ exchanger, NCX2 and NCX3, show a deletion of 37 amino acids in the intracellular loop corresponding to parts of the variable region of NCX1. We identified three splicing isoforms of NCX3 in brain and skeletal muscle by reverse transcriptase-polymerase chain reaction. These splice variants are generated by including either of two alternative exons equivalent to the NCX1 exon A or B and by including or excluding a sequence equivalent to the NCX1 exon C. We did not detect any alternative splicing of NCX2. We examined selected tissues from neonatal and adult rats and found developmental regulation for NCX1 and NCX3 splicing isoforms in skeletal muscle. Specific isoform patterns were also detected for NCX1 and NCX3 in cultured cortical neurons, astrocytes, and oligodendrocytes. We suggest a new terminology to distinguish the different splice variants of individual NCX isoforms.

Differentiation of Naegleria fowleri from other species of Naegleria using monoclonal antibodies and the polymerase chain reaction

1994 ◽  
Vol 88 (1) ◽  
pp. 119-120 ◽  
Author(s):  
Olivier Sparagano ◽  
Emmanuel Drouet ◽  
Gérard Denoyel ◽  
Pierre Pernin ◽  
Marie-Hélène Ruchaud-Sparagano
Keyword(s):  
Polymerase Chain Reaction ◽  
Monoclonal Antibodies ◽  
Naegleria Fowleri ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Nægleria Fowleri

scholarly journals Full-Length Sequence of Mouse Acupuncture-Induced 1-L (Aig1l) Gene Including Its Transcriptional Start Site

2011 ◽  
Vol 2011 ◽  
pp. 1-7
Author(s):  
Mika Ohta ◽  
Aki Sugano ◽  
Shuji Goto ◽  
Surini Yusoff ◽  
Yushi Hirota ◽  
...  
Keyword(s):  
Gene Expression ◽  
Skeletal Muscle ◽  
Polymerase Chain Reaction ◽  
Tissue Distribution ◽  
Cdna Sequence ◽  
Transcriptional Start Site ◽  
Full Length ◽  
Start Site ◽  
Chain Reaction ◽  
Polymerase Chain

We have been investigating the molecular efficacy of electroacupuncture (EA), which is one type of acupuncture therapy. In our previous molecular biological study of acupuncture, we found an EA-induced gene, named acupuncture-induced 1-L (Aig1l), in mouse skeletal muscle. The aims of this study consisted of identification of the full-length cDNA sequence ofAig1lincluding the transcriptional start site, determination of the tissue distribution ofAig1land analysis of the effect of EA onAig1lgene expression. We determined the complete cDNA sequence including the transcriptional start site via cDNA cloning with the cap site hunting method. We then analyzed the tissue distribution ofAig1lby means of northern blot analysis and real-time quantitative polymerase chain reaction. We used the semiquantitative reverse transcriptase-polymerase chain reaction to examine the effect of EA onAig1lgene expression. Our results showed that the complete cDNA sequence ofAig1lwas 6073 bp long, and the putative protein consisted of 962 amino acids. All seven tissues that we analyzed expressed theAig1lgene. In skeletal muscle, EA induced expression of theAig1lgene, with high expression observed after 3 hours of EA. Our findings thus suggest that theAig1lgene may play a key role in the molecular mechanisms of EA efficacy.

scholarly journals A review of Cryptosporidium spp. and their detection in water

2020 ◽  
Author(s):  
Eman M. Hassan ◽  
Banu Örmeci ◽  
Maria C. DeRosa ◽  
Brent R. Dixon ◽  
Syed A. Sattar ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Monoclonal Antibodies ◽  
Sensitivity And Specificity ◽  
Gastrointestinal Diseases ◽  
Dna Aptamers ◽  
Chain Reaction ◽  
Cryptosporidium Spp ◽  
Water Matrix ◽  
Polymerase Chain ◽  
Near Future

Abstract Cryptosporidium spp. are one of the most important waterborne pathogens worldwide and a leading cause of mortality from waterborne gastrointestinal diseases. Detection of Cryptosporidium spp. in water can be very challenging due to their low numbers and the complexity of the water matrix. This review describes the biology of Cryptosporidium spp. and current methods used in their detection with a focus on C. parvum and C. hominis. Among the methods discussed and compared are microscopy, immunology-based methods using monoclonal antibodies, molecular methods including PCR (polymerase chain reaction)-based assays, and emerging aptamer-based methods. These methods have different capabilities and limitations, but one common challenge is the need for better sensitivity and specificity, particularly in the presence of contaminants. The application of DNA aptamers in the detection of Cryptosporidium spp. oocysts shows promise in overcoming these challenges, and there will likely be significant developments in aptamer-based sensors in the near future.

GRIM19 is involved in WT1 expression and epithelial-to-mesenchymal transition in adenomyotic lesions

Reproduction ◽  
2021 ◽  
Author(s):  
Chen Geng ◽  
Hao-ran Liu ◽  
Yue Zhao ◽  
Yang Yang ◽  
Lan Chao
Keyword(s):  
Polymerase Chain Reaction ◽  
Real Time ◽  
Western Blotting ◽  
Small Interfering Rna ◽  
Epithelial To Mesenchymal Transition ◽  
Chain Reaction ◽  
Mesenchymal Transition ◽  
Ishikawa Cells ◽  
Polymerase Chain ◽  
Interfering Rna

The epithelial-to-mesenchymal transition may play a role in adenomyosis. GRIM19 expression is downregulated in adenomyotic lesions, and the effects of this downregulation in adenomyosis remain relatively unclear. We aimed to explore whether aberrant GRIM19 expression is associated with the epithelial-to-mesenchymal transition in adenomyosis. In this study, expression of both GRIM19 and WT1 was low, and epithelial-to-mesenchymal transition, which included significant changes in CDH1, CDH2 and KRT8 expression, occurred in adenomyotic lesions, as confirmed by western blotting and quantitative real-time polymerase chain reaction. We provided novel insights into WT1 expression in adenomyosis, revealing that WT1 expression was increased in the endometrial glands of adenomyotic lesions by immunohistochemistry. In vitro, knockdown of GRIM19 expression by small-interfering RNA promoted the proliferation, migration and invasion of Ishikawa cells, as measured by Cell Counting Kit-8, wound healing assay and Transwell assays. Western blotting and quantitative real-time polymerase chain reaction confirmed that WT1 expression increased and epithelial-to-mesenchymal transition was induced, including upregulation of CDH2 and downregulation of CDH1 and KRT8 after transfecting the GRIM19 small interfering RNA to Ishikawa cells. Furthermore, WT1 expression was upregulated and epithelial-to-mesenchymal transition was observed, including downregulation of CDH1 and KRT8 in GRIM19 gene-knockdown mice. Upregulation of WT1 expression in the endometrial glands of GRIM19 knockdown mice was also verified by immunohistochemistry. Taken together, these results reveal that low expression of GRIM19 in adenomyosis may upregulate WT1 expression and induce epithelial-to-mesenchymal transition in the endometria, providing new insights into the pathogenesis of adenomyosis.

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