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Updated: Jun 27, 2026
High-throughput Detection Method for Influenza Virus
Published on: February 4, 2012
Immunofluorescence imaging of the influenza virus M1 protein is dependent on the fixation method
Toshikatsu Shibata1, Torahiko Tanaka, Kazufumi Shimizu
1Division of Microbiology, Department of Pathology and Microbiology, Nihon University School of Medicine, 30-1 Oyaguchi-kami-cho, Itabashi-ku, Tokyo, 173-8610, Japan.
Insights
Fixation method significantly impacts influenza virus M1 protein localization. Methanol fixation reveals nuclear dots of M1 protein, co-localized with PML protein, unlike paraformaldehyde fixation.
Area of Science:
- Virology
- Cell Biology
- Immunofluorescence Microscopy
Background:
- The matrix (M1) protein is crucial for influenza virus assembly and replication.
- Understanding M1 protein distribution within infected cells is key to elucidating viral mechanisms.
- Immunostaining techniques are vital for visualizing protein localization in cellular contexts.
Purpose of the Study:
- To investigate the subcellular distribution of the influenza virus M1 protein in infected cells.
- To determine the influence of different fixation methods on M1 protein immunofluorescence patterns.
- To assess the temporal appearance and nuclear localization of M1 protein.
Main Methods:
- Immunostaining of influenza virus-infected cells using specific antibodies against M1 protein.
- Comparison of fixation methods: paraformaldehyde versus methanol.
- Time-course analysis of M1 protein distribution post-inoculation.
- Co-localization studies with promyelocytic leukemia (PML) protein.
Main Results:
- Paraformaldehyde fixation resulted in uniform M1 protein distribution in cytoplasm and nuclei.
- Methanol fixation clearly revealed distinct nuclear dots of M1 protein, visible from 8 hours post-inoculation.
- Diffuse M1 protein distribution was observed up to 6 hours post-inoculation.
- Nuclear M1 protein dots co-localized with PML protein, a component of nuclear domain 10 (ND10).
Conclusions:
- The observed nuclear dots of M1 protein are not fixation artifacts but represent a genuine cellular localization.
- Methanol fixation is the preferred method for accurate visualization and localization of influenza M1 protein within the nucleus.
- The findings provide insights into the dynamic nuclear behavior of influenza M1 protein during infection.
Abstract:
The distribution of the matrix (M1) protein of influenza virus in infected cells was examined using immunostaining. The fixation method influenced strongly the immunofluorescence pattern of the M1 protein. The M1 protein was distributed uniformly in both the cytoplasm and in nuclei when cells that had been infected with virus were fixed with paraformaldehyde. In cells that had been fixed with methanol, however, nuclear dots of the M1 protein were clearly visible. The dots were evident at 8h post-inoculation. Up to 6h post-inoculation, only a diffuse distribution of the M1 protein was observed. The dots were co-localized with promyelocytic leukemia (PML) protein, a major component of nuclear domain 10 (ND10), also called PML oncogenic domains (PODs) or PML-nuclear bodies (NBs). These results indicate that the nuclear dots of the M1 protein in cells that had been fixed with methanol are not artifacts of the fixation method. Furthermore, methanol fixation is preferred for localization of the influenza M1 protein in nuclei using immunostaining.
