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Quantitative immunocytochemistry using an image analyzer. II. Concentration standards for transmitter
L B Nabors1, E Songu-Mize, R R Mize
1Department of Anatomy and Neurobiology, College of Medicine, University of Tennessee Health Science Center, Memphis 38163.
Journal of Neuroscience Methods
|November 1, 1988
Summary
This study introduces a method to quantify neurotransmitter concentration using immunocytochemistry and image analysis. The technique establishes a linear relationship between optical density and neurotransmitter levels, enabling accurate measurement in biological samples.
Area of Science:
- Neuroscience
- Biochemistry
- Immunology
Background:
- Image analyzers can quantify immunocytochemistry labeling.
- Estimating neurotransmitter concentration is crucial for neuroscience research.
- Previous methods may lack precision or require complex procedures.
Purpose of the Study:
- To develop and validate a method for estimating neurotransmitter concentration using immunocytochemistry and optical density measurements.
- To create a reliable standard for quantifying neurotransmitters in artificial media.
Main Methods:
- Developed a standard using a 3% agar matrix activated with cyanogen bromide/acetonitrile.
- Used gamma-aminobutyric acid (GABA) conjugated to bovine serum albumin (BSA) as the antigen.
- Incubated activated agar with serial dilutions of labeled GABA/BSA conjugate, followed by antibody incubation and immunocytochemistry.
- Measured optical density using an image analyzer.
Main Results:
- A 3% agar matrix proved optimal for standard preparation.
- A linear relationship was established between GABA concentration and optical density (0.01 to 1 nmol/mg agar).
- The method accurately estimates bound GABA concentration from immunocytochemical labeling.
Conclusions:
- The developed immunocytochemistry-based method allows for reliable estimation of neurotransmitter concentration.
- This technique offers a valuable tool for quantifying neurotransmitters in biological tissues.
- Further applicability to fixed brain tissue is discussed.