Comparative Analysis of MBNL1 Antibodies: Characterization of Recognition Sites and Detection of RNA Foci Colocalization
Abstract
:1. Introduction
2. Materials and Methods
2.1. Plasmid Construction
2.2. Cell Culture
2.3. Transfection
2.4. Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE)
2.5. Reverse Transcription-PCR (RT-PCR)
2.6. Fluorescence In Situ Hybridization (FISH)
2.7. Immunofluorescence Staining
2.8. Immunoprecipitation
3. Results
3.1. Differential Detection of MBNL1 by Various Antibodies
3.2. Mapping of MBNL1 Antibody Recognition Sites Using Deletion Mutants
3.3. Comparison of Antibody Reactivity to MBNL Paralogs
3.4. Endogenous Expression Patterns of MBNL1
3.5. Detection of RNA Foci
3.6. Detection of MBNL1 by Immunoprecipitation
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
DM | Myotonic dystrophy |
DM1 | Myotonic dystrophy type 1 |
DM2 | Myotonic dystrophy type 2 |
EGFP | Enhanced green fluorescent protein |
FISH | Fluorescence In Situ Hybridization |
IF | Immunofluorescence |
MBNL1 | Muscleblind-like 1 |
ZnF | Zinc finger |
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MBNL1 Antibody | Company | Catalog Number | Host Species | Immunogen |
---|---|---|---|---|
3E7 | Abnova (Taipei, Taiwan) | H00004154-M02 | Mouse monoclonal | Full-length recombinant human MBNL1 |
4A8 | Santa Cruz Biotechnology (Dallas, TX, USA) | sc-136165 | Mouse monoclonal | Full-length recombinant human MBNL1 |
MBNL1/2/3 | Santa Cruz Biotechnology | sc-58790 | Mouse monoclonal | A 371 amino acid fragment of human MBNL1 |
66837-1-Ig | Proteintech (Rosemont, IL, USA) | 66837-1-Ig | Mouse monoclonal | Recombinant protein (amino acids 59–400 of human MBNL143) |
ab45899 | Abcam (Cambridge, UK) | ab45899 | Rabbit polyclonal | Synthetic peptide (amino acids 250–350 of human MBNL1) |
ARP41227 | Aviva Systems Biology (San Diego, CA, USA) | ARP41227_P050 | Rabbit polyclonal | Synthetic peptide (amino acids 268–317) |
MBNL1 Antibody | Recognition Site | RNA Foci Detection | Immunoprecipitation |
---|---|---|---|
3E7 | 139–161 (exon 3) | Applicable | Applicable |
4A8 | 139–161 (exon 3) | Applicable | Applicable |
sc-58790 | 184–238 (exon 4) | Not applicable | Not applicable |
66837-1-Ig | 139–161 (exon 3) | Not applicable | Applicable |
ab45899 | 299–308 (exon 6) | Applicable | Applicable (recommended) |
ARP41227 | 270–275 (exon 5) | Applicable | Applicable (not efficient) |
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Aoki, Y.; Ohki, A.; Yanaizu, M.; Kino, Y. Comparative Analysis of MBNL1 Antibodies: Characterization of Recognition Sites and Detection of RNA Foci Colocalization. Genes 2025, 16, 658. https://doi.org/10.3390/genes16060658
Aoki Y, Ohki A, Yanaizu M, Kino Y. Comparative Analysis of MBNL1 Antibodies: Characterization of Recognition Sites and Detection of RNA Foci Colocalization. Genes. 2025; 16(6):658. https://doi.org/10.3390/genes16060658
Chicago/Turabian StyleAoki, Yoshitaka, Ai Ohki, Motoaki Yanaizu, and Yoshihiro Kino. 2025. "Comparative Analysis of MBNL1 Antibodies: Characterization of Recognition Sites and Detection of RNA Foci Colocalization" Genes 16, no. 6: 658. https://doi.org/10.3390/genes16060658
APA StyleAoki, Y., Ohki, A., Yanaizu, M., & Kino, Y. (2025). Comparative Analysis of MBNL1 Antibodies: Characterization of Recognition Sites and Detection of RNA Foci Colocalization. Genes, 16(6), 658. https://doi.org/10.3390/genes16060658