Description
Hematoxylin is oxidized to hematein, which is complexed with aluminum mordant to form a dye-metal complex. This complex selectively binds to nuclear chromatin and nucleic acids. Following rinsing and bluing, cell nuclei develop a permanent blue to blue-purple coloration while cytoplasmic structures remain available for counterstaining.
Gill Hematoxylin II is formulated as a progressive stain, allowing the desired staining intensity to be controlled primarily by immersion time rather than by differentiation.
VitroView™ Gill Hematoxylin II Solution is a ready-to-use, double-strength progressive nuclear stain designed for use in histology, cytology, and immunohistochemistry (IHC). The formulation produces well-defined blue to blue-purple nuclear staining with minimal background and is suitable for routine Hematoxylin and Eosin (H&E) staining of paraffin-embedded tissues.
Gill Hematoxylin II provides greater staining intensity than Gill Hematoxylin I while requiring little or no differentiation under standard progressive staining protocols.
Application
- Progressive nuclear staining of formalin-fixed, paraffin-embedded (FFPE) tissue sections
- Routine Hematoxylin and Eosin (H&E) staining
- Cytology staining where increased nuclear intensity is desired
- Counterstaining in immunohistochemistry (IHC)
Kit Contents
VB-3050G2 VitroView™ Gill Hematoxylin II Solution —– 500 ml
Storage Condition
Room temperature.
Protocol
H&E Stain for Paraffin Sections
- Deparaffinize in xylene I for 6 minutes and II for 6 minutes.
- Rehydrate: ethanol 100% (2 minutes)x2; ethanol 95% (2 minutes)x2; ethanol 70% (2 minutes)x1; Rinse in distilled water (5 minutes).
- Place in Gill Hematoxylin II Solution for 2-4 minutes (adjust according to tissue type and desired intensity).
- Rinse in tap water for 5 min with 3 changes. Or Bluing reagent for 30–60 seconds.
- Rinse with water for 1 min.
- Place in 95% Ethanol for 1 minute (DO NOT RINSE).
- Place in Eosin for 1 min 30sec.
- Dehydrate with 2 changes of 95% Ethanol and 2 changes of 100% Ethanol (2 minutes per change).
- Clear with 3 changes of xylene (5 minutes per change)
- Mount coverslip onto glass slide with Permount or some other suitable organic mounting medium.
H&E Stain for Frozen Sections
- Frozen section are fixed in acetone at -20 °C for 3 min or 80% Methanol at 4°C for 5 min
- PBS x 10 min.
- Rinse in DI water (2 minutes).
- Follow step 3-10 above.
For Immunohistochemistry (IHC) Counterstain:
Following completion of chromogen development:
- Rinse slides thoroughly.
- Immerse in Gill Hematoxylin I for 30–90 seconds.
- Rinse with water.
- Blue nuclei.
- Dehydrate, clear, and mount.
Counterstaining time may be adjusted depending on tissue type and desired nuclear intensity.
Note: This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.
Precautions: Handle with care. Avoid contact with eyes, skin and clothing. Do not ingest. Wear gloves.
