Movat Pentochrome
A histological stain used to differentiate connective tissue components, highlighting collagen, muscle, fibrin, mucin, and elastic fibers in tissue samples.
Author: Kelsey Goldman, HTL (ASCP)
Last Update: 1 March 2025
Copyright: 2025, HistologyOutlines.com
Cite this Page! -> Goldman, Kelsey. “Movart Pentochrome.” HistologyOutlines.Com, 1 March 2025. Accessed (today).
Purpose
- To clearly demonstarate mucin, fibrin, elastic fibers, muscle, collagen in tissue (Carson, 2001).
Control Tissues
- Lung, Skin, Colon (Carson, 2001)
Staining Pattern
- Nuclei and Elastic Fibers -Black
- Collagen- Yellow
- Ground Substance/ Mucins- Blue
- Fibrins- Intense Red
- Muscle- Red
Biochemistry Theory
- Alcian blue- stains acidic muco-substances; the Alcian blue is post stained by alkaline alcohol, which makes the stained molecules insoluble. the stained product here is called monastral fast blue. the unconverted alcian blue will be lost as the staining progresses. The mucin and ground substance will be stained blue(Carson, 2001).
- Elastic Fibers are stained by iron hematoxylin that is differentiated by a ferric chloride solution through the Veroeff staining method. This method uses iodine, therefore sodium thiosulfate must be used after this staining step in order to remove excess coloration from the ioodine. The elastic fibers will be stained black(Carson, 2001).
- Crocein scarlet and acid fusion are used to stain muscles, cytoplasm, collagen and the ground substances. with the addition of phosphortungstic acid a differentiation occurs in which staining is removed from the ground substance and the collagen. Leaving only the muscle and cytoplasm stained red at this step in staining. Phosphortungstic acid is removed through the adding of acetic acid(Carson, 2001).
- Alcoholic Safran is used to counterstain the collagen yellow(Carson, 2001).
Troubleshooting
- Ensure that the fixative used is either 10% NBF or acetic formalin sublimate(Carson, 2001).
Additional References
- Carson, F. L. (2001). Histotechnology: a self-instructional text (2. ed., [Reprint]). ASCP Press.
Sample Protocol
- Alcian Blue 1%
- Alkaline alcohol
- 10mL ammonium hydroxide
- 90 mL 95% Alcohol
- Iodine Iodide
- 2g Iodine
- 4g Potassium Iodide
- 100 mL diH2O (add the reagents to 25mL of water initially, mix, then add the remaining water)
- Absolute Alcoholic Hematoxylin 10% (make ahead and store at room temp)
- Hematoxylin 10g
- 100mL Absolute Alcohol
- Ferric chloride 10% (make ahead and store at RT)
- 10g Ferric chloride
- 100 mL H2O
- Hematoxylin solution (make fresh)
- Combine 25 mL each of the absolute alcoholic hematoxylin, absolute alcohol, 10% ferric chloride, and iodine-iodide.
- Ferric Chloride 2%
- 10% Ferric chloride 10mL
- 40 mL diH2O
- 5% Sodium thiosulfate
- Crocein Scarlet-Acid Fuchsin
- Solution A (stock, store at RT)
- Crocein Scarlet 1g
- diH2O 99.5 mL
- Glacial Acetic Acid 0.5 mL
- Solution B (stock, store at RT)
- Acid fuchsin 0.1g
- diH2O 99.5 mL
- Glacial Acetic Acid 0.5 mL
- Working solution (make fresh)
- 8 parts Solution A
- 2 parts Solution B
- Solution A (stock, store at RT)
- Phosphotungstic Acid 5%
- Alcoholic Safran
- Safran du Gatinais 6g
- Absolute alcohol 100 mL
- Deparaffinize and bring to water
- Place slides into Alcian Blue for 20 minutes
- Wash in running tap water for 5 minutes
- Place slides into a jar of alkaline alcohol for 1 hour
- Wash in running tap water for 10 minutes. Do not underperform this step. If alkaline alcohol is left behind, the rest of the staining will not yield correct results.
- Quick rinse in diH2O
- Stain in the freshly made hematoxylin solution for 15 minutes
- Rinse in several changes of diH2O
- Place in 2% ferric chloride to differentiate. Differentiation is complete when the elastic fibers appear sharp from the rest of the background when viewed under the microscope. This process will be complete in 2-3 minutes.
- Rinse in diH2O
- Place into 5% sodium thiosulfate for 1 minute
- Wash in running tap water for 5 minutes followed by a quick rinse in diH2O
- Stain in the crocein scarlet-acid fuchsin for 1.5 minutes
- Run through several changes of diH2O
- Rinse in 0.5% acetic acid solution
- Stain slides in 5% phosphotungstic acid solution. Use two Coplin jars of identical solution, stain the slides in each jar for 5 minutes each.
- Rinse the slides in 0.5% acetic acid solution
- Rinse the slides in three changes of absolute alcohol
- Counterstain in alcoholic Safran solution for 15 minutes
- Rinse the slides in three changes of absolute alcohol
- Dehydrate and run through xylene, coverslip with a xylene-based mounting media.
Reagents
Method
- Adapted from (Carson, 2001)