Gomori Aldehide Fuschin Staining for Pancreatic Beta Cell and Elastic Fiber

Protocol After Paraffin Block Sectioning

1. Rehydration (Xylene 2x5 minutes, ethanol 100% 2 minutes, ethanol 95% 2x2 minutes, ethanol 70% 2 minutes, deionized water 2 minutes) 
2. Gomori Aldehide Fuschin Staining 1 hours then wash deionized water 10 dips
3. Nuclear Fast Red for 5 minutes then wash in tap water for 5 minutes  
4. Dehydration (ethanol 70% 10 seconds, ethanol 95% 20 seconds, ethanol 100% 10 seconds, xylene 2x10 seconds) 
5. Mounting in Xylene based medium

Result

Recepie for Gomori Aldehide Fuschin:

1. Weight 2.4 gr picric acid solid to a bronze bottle, added with water to 200ml, shaking for 5-10mins (Picric acid stored in 36% water, so pulled out water before weighting, remember to return back the water to the bottle after weighting)
2. Add 0.2 gr Direct Red 80 to above solution, then shaking/mixing for 5 minutes

Recepie for Nuclear Fast Red:

1. Weight 2.4 gr picric acid solid to a bronze bottle, added with water to 200ml, shaking for 5-10mins (Picric acid stored in 36% water, so pulled out water before weighting, remember to return back the water to the bottle after weighting)
2. Add 0.2 gr Direct Red 80 to above solution, then shaking/mixing for 5 minutes

Recommended Reagents:

1. Paraldehyde (Sigma Aldrich) 
2. 
   
3. Nuclear Fast Red (Sigma Aldrich)

Collagen Staining by Picro-Sirius Red

Protocol After Paraffin Block Sectioning

1. Rehydration (Xylene 2x5 minutes, ethanol 100% 2 minutes, ethanol 95% 2x2 minutes, ethanol 70% 2 minutes, deionized water 2 minutes) 
2. Picro-Sirius Red staining 1 hours then wash deionized water 10 dips
3. Gill Hematoxylin for 5 minutes then wash in tap water for 5 minutes  
4. Dehydration (ethanol 70% 10 seconds, ethanol 95% 20 seconds, ethanol 100% 10 seconds, xylene 2x10 seconds) 
5. Mounting in Xylene based medium

Result

Recepie for Picro-Sirius Red:

1. Weight 2.4 gr picric acid solid to a bronze bottle, added with water to 200ml, shaking for 5-10mins (Picric acid stored in 36% water, so pulled out water before weighting, remember to return back the water to the bottle after weighting)
2. Add 0.2 gr Direct Red 80 to above solution, then shaking/mixing for 5 minutes

Recommended Reagents:

1. Picric acid (Sigma Aldrich) NOTE: Becareful picric acid is toxic and potentially explosive when dry)
2. Direct Red 80 (Sigma Aldrich)

Detection of MMP1 by Immunohistochemistry in Cancer, Aging and Rheumatoid Arthritis

MMPs or matrix metalloproteinases are member of peptidase. These enzymes have capability for degradation extracellular matrix components (collagen, gelatin, Fibronectin, Laminin and proteoglycan. MMP genes can be activated by phorbol ester, lipopolysaccharide (LPS) or staphylococcal enterotoxin B (SEB). For doing its job, MMP requires both calcium and zinc. MMP-1 is downregulated by p53, and abnormality of p53 expression may contribute to joint degradation in rheumatoid arthritis by regulating MMP-1 expression.

  

IHC of MMP1 in Mouse Lung (Abbiotec)


Western blott Analysis of Human MMP1(Santa Cruz)

Principal of Protocol: 

1. Rehydration (Xylene 2x5 minutes, ethanol 100% 2 minutes, ethanol 95% 2x2 minutes, ethanol 70% 2 minutes, PBS 2 minutes) 
2. Antigen Retrieval (Citrate buffer in 800 Watt Microwave for 5 minutes) then wash PBS 2x5 minutes 
3. Blocking for endogenous peroxidase (Hydrogen Peroxide 3% for 5 minutes)then wash PBS 2x5 minutes 
4. Primary Antibody (1 hours in room temperature 25 degree Celcius)then PBS 2x5 minutes 
5. Secondary Antibody (Depend on manufacture, in this case using LSAB+ Kit Dako), then wash 2x5 minutes 
6. Dehydration (ethanol 70% 10 seconds, ethanol 95% 20 seconds, ethanol 100% 10 seconds, xylene 2x10 seconds) 
7. Mounting in Xylene based medium 

Recommended Reagent:

1. Citrate Buffer (Trisodium Citrate Sigma Aldrich) 
2. Primary Antibody (Santa Cruz, Abbiotec) 
3. Secondary antibody LSAB+ kit (Dako) 
4. Xylene or substitute (Merck, Shandon) 

Citation:
Santa Cruz
Abbiotec
DAKO