Table of Contents
What is tissue autofluorescence?
Autofluorescence is a general term describing the background fluorescence in tissue sections unrelated to the specific signal generated during an IF assay. Tissue components such as red blood cells (RBCs) and collagen are strongly fluorescent, making it difficult to discern between relevant signal and background.
What is the function of autofluorescence?
Fundus autofluorescence (FAF) is an adjunctive tool for the assessment of CSC. During the acute phase of the disease, FAF typically shows hypofluorescence at the leakage point and over the area of neurosensory detachment due to blockage by subretinal fluid.
What causes cell autofluorescence?
Cellular autofluorescence can be due to the presence of collagen and elastin, cyclic ring compounds such as NADPH and riboflavin, aromatic amino acids and cellular organelles such as mitochondria and lysosomes.
What is autofluorescence in flow cytometry?
MIT Flow Cytometry Core Facility. Page 2. Autofluorescence. Cells contain molecules, which become fluorescent when excited by UV/Visual radiation of suitable wavelength. This fluorescence emission, arising from endogenous fluorophores, is an intrinsic property of cells and is called auto-fluorescence.
Where does autofluorescence come from?
Autofluorescence frequently comes not from the sample, but from the way it is treated prior to imaging. For example, mounting media, tissue culture media and laboratory plastic can all be sources of autofluorescence.
What is fundus autofluorescence test?
Fundus autofluorescence (FAF) is a non-invasive imaging modality that has become increasingly popular in both research and clinical settings due to its ability to map naturally and pathologically occurring fluorophores in the posterior pole.
Do autofluorescence do dead cells?
Dead cells can bind non-specifically with a lot of reagents, increase autofluorescence significantly, and alter scatter properties. The presence of extracellular matrix debris also contributes to autofluorescence through collagen and elastin.
Does collagen have autofluorescence?
Collagen is highly autofluorescent. The best way to get rid of it is to subtract fluorescence images prior to adding the antibody from images take after staining with the antibody.
How do you deal with autofluorescence?
5 Tips to Reduce Autofluorescence
- Use a lower concentration of FCS in the staining buffer.
- Remove dead cells & other debris.
- Lyse RBCs properly & remove lysed contents.
- Lower PFA concentration & avoid storing cells in PFA for long durations.
- Make the right fluorochrome choices.
What does autofluorescence look like?
Normal Fundus Autofluorescence In a normal fundus without retinal pathology, blood vessels will appear dark since blood is able to strongly absorb the blue (~488nm) or green (~514nm) light that is typically used in FAF imaging.
How do I remove autofluorescence from tissue?
Historically, the main method that has been employed to lower tissue autofluorescence has been to treat the tissue with solutions of Sudan Black or similar nonfluorescent diazo dyes. These hydrophobic dye molecules will generally bind nonspecifically to tissue sections.
How is fundus autofluorescence imaging performed?
Unlike fluorescein angiography, FAF does not require the injection of a fluorescein dye in order to image the retina, but rather utilizes the fluorescent properties of lipofuscin within the retinal pigment epithelium (RPE) to create an image.
Why is autofluorescence a problem in fluorescence microscopy?
Autofluorescence exhibited by cells and tissues in culture can often limit the ability to detect fluorescent probes in stained and fixed preparations.
What is an autofluorescence test?
Fundus autofluorescence is a non-invasive diagnostic test that involves taking digital photographs of the back of the eye without a contrast dye.
What is an OCT used for?
Optical coherence tomography, or OCT, is an imaging method used to generate a picture of the back of your eye, called your retina. The noninvasive method produces an image by measuring the amount of a dim red light that reflects off of your retina and optic nerve.
What are the advantages of fluorescence microscopy?
Fluorescence microscopy is closely allied to transmission (absorption) microscopy in its range of application, but possesses particular advantages: great sensitivity for detection and quantification of small amounts of fluorescent substances or small particles, and the possibility of application to opaque objects.
What does fundus autofluorescence show?
Fundus autofluorescence (FAF), a relatively new imaging modality, focuses on the fluorescent properties of pigments in the retina to generate images that help us view various disease processes from a different perspective. It aids us in the understanding of the pathophysiology of different retinal disorders.
What is autofluorescence bronchoscopy?
Autofluorescence bronchoscopy is a bronchoscopic procedure in which a blue light rather than a white light is employed for illumination, and premalignant and malignant tissue is distinguished by a change in color from normal tissue without the need for fluorescence-enhancing drugs.
What is autofluorescence in microbiology?
The term “autofluorescence” is used to distinguish the intrinsic fluorescence of cells and tissues from the fluorescence obtained by treating specimens with exogenous fluorescent markers that binds cell and tissue structures.
What are the advantages of auto-fluorescence?
Therefore, analytical techniques based on auto-fluorescence monitoring can be utilized in order to obtain information about morphological and physiological state of cells and tissues. Moreover, auto-fluorescence analysis can be performed in real time because it does not require any treatment of fixing or staining of the specimens.
What is the role of autofluorescence in the detection of arterial plaque?
Laser-induced autofluorescence has been used to detect calcified and noncalcified plaques in arterial tissues. Autofluorescence signals from normal artery correlate with the structural proteins elastin and collagen present in intima, media and adventitia.
What is the purpose of the autofluorescence report?
Our report is intended to confirm the reference and quantitate the autofluorescence with reference to individual patients.