Immunofluorescence microscopy
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Immunofluorescence Microscopy PowerPoint PPT Presentation


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Immunofluorescence Microscopy. Immunofluorescence Microscopy:. When an antibody, or the antiimmunoglobulin antibody used to detect the antibody is labeled with a fluorescent dye This method is used when looking at the subcellular location of a protein of interest.

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Immunofluorescence Microscopy

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Immunofluorescence microscopy

ImmunofluorescenceMicroscopy


Immunofluorescence microscopy1

Immunofluorescence Microscopy:

  • When an antibody, or the antiimmunoglobulin antibody used to detect the antibody is labeled with a fluorescent dye

  • This method is used when looking at the subcellular location of a protein of interest


Immunofluorescence microscopy2

Immunofluorescence Microscopy:

  • See Page 162 in textbook for another diagram


Technique

Technique:

  • Common dyes: fluorescein, rhodamine

  • Dyes chosen are excited by a certain light wavelength, usually blue or green, and emit light of a different wavelength in the visible spectrum

    • Eg. Fluorescein emits green light

    • Eg. Rhodamine emits orange/red light

  • By using selective filters in a fluorescence microscope only the light from the dye is detected

  • Available fluorescent labels now include red, blue, cyan or yellow fluorescent proteins


Immunofluorescence microscopy

Uses:

  • This can be used to detect the distribution of any protein

  • By attaching different dyes to different antibodies the distribution of two or more molecules can be determined in the same cell or tissue sample


Technology

Technology:

  • Confocal fluorescent microscope: uses a computer-aided techniques to produce a extremely thin optical section of a cell or tissue

    • Gives high resolution microscopy without elaborate sample preparation


The zeiss lsm 410 confocal microscope

The Zeiss LSM 410 confocal microscope


Technology continued

Technology Continued:

  • Time-lapse video microscopy: sensitive digital video cameras record the movement of fluorescently labeled molecules in cell membranes and their redistribution when cells come into contact with one another


References

References:

  • Class textbook (Human Molecular Genetics 3)

  • www.antibodystation.com/immunofluorescence-microscopy/

  • National University of Singapore website www.med.nus.edu.sg/path/services/immunofluor.htm

  • BD Biosciences website www.bdbiosciences.com


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