Skip to product information
Request a Quote
Crystal Violet Powder | Gram Stain Dye for Microbiology
$37.00 AUD
Documentation
Crystal Violet
Biological Staining Dye — Gram Staining & Laboratory Research | CAS 548-62-9
Cat. No. ASD-606 Basic Violet 3 | Gentian Violet C₂₅H₃₀ClN₃ MW 407.98 g/mol 🇦🇺 Made in Melbourne ⚡ Same-Week Dispatch
Overview & Biochemistry
Crystal Violet (Basic Violet 3) is a cationic triphenylmethane dye and one of the most versatile staining reagents in the microbiology and histology laboratory. Its intense purple coloration and strong affinity for negatively charged cellular components — particularly the peptidoglycan layer of bacterial cell walls and nucleic acids — make it the primary stain in the Gram staining procedure and a standard reagent across a wide range of cell-based assays.
The dye is also known as Gentian Violet in clinical contexts, where it has historically been used as a topical antimicrobial agent, and as Methyl Violet 10B in industrial dye classifications.
Mode of Action
Cationic Binding → Cell Wall Penetration → Iodine Mordanting → Differential Retention
Crystal Violet is a positively charged dye at laboratory pH. It penetrates the cell wall and membrane of both Gram-positive and Gram-negative bacteria and forms a dye–iodine complex when treated with Gram's iodine (mordant). In Gram-positive bacteria, the thick peptidoglycan layer contracts during decolourisation with acetone-alcohol, trapping the CV–iodine complex and retaining the purple coloration. In Gram-negative bacteria, the thin peptidoglycan and outer lipopolysaccharide membrane are disrupted by the decolourant, releasing the complex — these cells are then counterstained with safranin (pink/red).
Crystal Violet is a positively charged dye at laboratory pH. It penetrates the cell wall and membrane of both Gram-positive and Gram-negative bacteria and forms a dye–iodine complex when treated with Gram's iodine (mordant). In Gram-positive bacteria, the thick peptidoglycan layer contracts during decolourisation with acetone-alcohol, trapping the CV–iodine complex and retaining the purple coloration. In Gram-negative bacteria, the thin peptidoglycan and outer lipopolysaccharide membrane are disrupted by the decolourant, releasing the complex — these cells are then counterstained with safranin (pink/red).
Applications
🧪
Gram Staining
Primary stain — Gram-positive identification
🔬
Microscopy
General cell & tissue staining
🧬
Biofilm Assays
CV crystal violet quantification
🪬
Cell Viability
Adherent cell biomass staining
📊
Colony Counting
Enhanced plate visualisation
🏫
Education
Teaching labs & demonstrations
Gram Staining Protocol — Step by Step
| 1 | Fix smear: Prepare a thin bacterial smear on a clean glass slide. Air dry, then heat-fix by passing through a flame 2–3 times. |
| 2 | Primary stain: Flood slide with Crystal Violet solution (ASD-606, 0.4–2% w/v in ethanol/water). Leave for 60 seconds. Gently rinse with distilled water. |
| 3 | Mordant: Apply Gram's Iodine solution. Leave for 60 seconds. Rinse gently with distilled water. The iodine forms an insoluble CV–iodine complex within cells. |
| 4 | Decolourise: Apply acetone-alcohol decolourant drop by drop until the solvent runs clear (typically 10–30 seconds). Immediately rinse with distilled water to stop decolourisation. This is the most critical step — over-decolourisation produces false Gram-negative results. |
| 5 | Counterstain: Apply Safranin O solution (ASD-613). Leave for 60 seconds. Rinse with distilled water. Blot dry — do not rub. |
| 6 | Read: Examine under oil immersion (×1000). Gram-positive = purple/violet. Gram-negative = pink/red. |
⚠️ Note: Decolourisation time is critical and varies with smear thickness, organism type, and decolourant concentration. Optimise for your specific protocol and organism. Old or thick smears may give false Gram-negative results.
Biofilm Quantification Assay (CV Method)
| 1 | Grow biofilm in microtitre plate wells under selected conditions. Remove planktonic cells by washing ×3 with PBS. |
| 2 | Add 0.1% Crystal Violet solution (100 µL/well). Stain for 15–20 minutes at room temperature. |
| 3 | Wash ×3 with distilled water to remove unbound dye. Air dry inverted for 30 minutes. |
| 4 | Solubilise bound CV with 200 µL 33% acetic acid per well for 10–15 minutes. |
| 5 | Read absorbance at 590 nm (OD₅₉₀). Higher OD = greater biofilm biomass. |
Cross-Reference / Equivalent Products
| Manufacturer | Product Name | Cat. No. | Notes |
|---|---|---|---|
| Sigma-Aldrich (Merck) | Crystal Violet | C3886 | Equivalent powder grade |
| Merck Millipore | Crystal Violet for microscopy (Certistain®) | 1.01408 | Equivalent powder grade |
| Thermo Fisher Scientific | Crystal Violet Solution | C3886 | Ready-to-use solution; ASD-606 is powder |
| Oxoid (Thermo Fisher) | Crystal Violet (Gram Staining Reagent) | BR0011 | Ready-to-use Gram stain reagent |
Cross-references are for convenience only. Catalogue numbers are trademarks of their respective owners.
Related Products
Chemical & Physical Properties
| Product Name | Crystal Violet |
| Catalogue Number | ASD-606 |
| Synonyms | Basic Violet 3; Gentian Violet; Methyl Violet 10B; Hexamethyl Pararosaniline Chloride |
| CAS Number | 548-62-9 |
| Colour Index | CI 42555 |
| Molecular Formula | C₂₅H₃₀ClN₃ |
| Molecular Weight | 407.98 g/mol |
| Grade | Biological Stain Grade |
| Appearance | Dark purple to green metallic-sheen crystalline powder |
| Purity | ≥90% dye content (biological stain titration, CI 42555) |
| Solubility | Soluble in water (~16 g/L at 25°C) and ethanol; slightly soluble in chloroform |
| λmax (absorption) | ~590 nm (aqueous solution) |
| Melting Point | ~215°C (with decomposition) |
| Charge | Cationic (positively charged at physiological pH) |
Quality Control Specifications
| Test Parameter | Specification |
|---|---|
| Appearance | Dark purple crystalline powder with metallic green sheen; free-flowing |
| Dye Content | ≥90% (CI 42555 titration) |
| Gram Staining Performance | Clear differentiation of Gram-positive (purple) from Gram-negative (colourless after decolourisation) cells |
| Solubility | Complete dissolution in distilled water at 10 mg/mL |
| λmax | 590 ± 5 nm (0.01% aqueous solution) |
| Batch COA | Available for every batch on request |
Working Solution Preparation
| Application | Concentration | Solvent | Notes |
|---|---|---|---|
| Gram Stain (primary) | 0.4–2.0% w/v | 20% ethanol in water | Standard Hucker's method |
| Biofilm assay (staining) | 0.1–0.5% w/v | Distilled water or methanol | 15–30 min at RT |
| General cell staining | 0.5–1.0% w/v | Distilled water | 2–5 min; rinse thoroughly |
| Colony visualisation | 0.05–0.1% w/v | Distilled water | Flood plate; rinse with water |
Storage & Stability
| Storage Temperature | 15–25°C (ambient, away from heat sources) |
| Container | Tightly sealed, light-resistant container |
| Protect From | Direct light, moisture, and strong oxidising agents |
| Shelf Life (powder) | 3 years from manufacture date when stored correctly |
| Working Solutions | Stable 12 months at RT in sealed dark bottle; filter before use if precipitate forms |
Literature & References
| # | Reference |
|---|---|
| 1 | Hucker GJ, Conn HJ. Methods of Gram Staining. New York State Agricultural Experiment Station Technical Bulletin. 1923;93:3–37. [Original Hucker's crystal violet formulation] |
| 2 | Murray PR, Baron EJ, Jorgensen JH, et al. (eds). Manual of Clinical Microbiology. 9th ed. ASM Press; 2007. [Gram stain procedure and troubleshooting] |
| 3 | O'Toole GA. Microtiter dish biofilm formation assay. Journal of Visualized Experiments. 2011;(47):e2437. [Crystal violet biofilm quantification method] |
| 4 | Christensen GD, Simpson WA, Bisno AL, Beachey EH. Adherence of slime-producing strains of Staphylococcus epidermidis to smooth surfaces. Infect Immun. 1982;37(1):318–326. [Seminal biofilm CV assay paper] |
| 5 | ISO 7218:2007. Microbiology of food and animal feeding stuffs — General requirements and guidance for microbiological examinations. [Gram stain as identification tool] |
📄 Full SDS available: Download the complete 16-section GHS Safety Data Sheet (Australian WHS Regulations 2023 / GHS 7th Edition) — contact support@ausamics.com.au
Section 1 — Identification
| Product Name | Crystal Violet |
| Catalogue No. | ASD-606 |
| Supplier | AuSaMicS Pty Ltd | ABN 56 676 640 467 |
| Address | 31 Longview CT, Thomastown VIC 3074, Australia |
| Emergency | Poisons Information Centre: 13 11 26 (24 hr, Australia) |
| Phone | +61 412 520 598 |
| support@ausamics.com.au |
Section 2 — Hazard Identification (GHS 7th Ed / WHS 2023)
| GHS Classification | Acute Tox. 4 (oral) — H302: Harmful if swallowed Acute Tox. 4 (dermal) — H312: Harmful in contact with skin Eye Irrit. 2 — H319: Causes serious eye irritation Muta. 2 — H341: Suspected of causing genetic defects Carc. 2 — H351: Suspected of causing cancer Aquatic Chronic 2 — H411: Toxic to aquatic life with long-lasting effects |
| Signal Word | WARNING |
| GHS Pictograms | ⚠️ Exclamation mark | ☣️ Health hazard | 🌿 Environmental hazard |
Sections 4–8 — First Aid, Fire & PPE
| Inhalation | Remove to fresh air. Seek medical attention if symptoms persist. Avoid generating dust. |
| Skin Contact | Remove contaminated clothing. Wash with soap and water ≥15 min. Note: staining of skin is expected and not hazardous; use disposable gloves. Seek advice if irritation develops. |
| Eye Contact | Irrigate with large amounts of water ≥15 min, hold eyelids apart. Seek immediate medical attention. |
| Ingestion | Do NOT induce vomiting. Rinse mouth. Call Poisons Centre 13 11 26. Seek medical attention immediately. |
| Extinguishing Media | Water spray, foam, dry powder, CO₂ |
| Respiratory Protection | P2 particulate filter respirator when weighing or handling bulk powder |
| Eye Protection | Safety glasses or chemical splash goggles — staining of eyes is a significant risk |
| Skin Protection | Nitrile gloves (≥0.11 mm), lab coat, closed footwear. CV stains skin and clothing permanently. |
Sections 12–15 — Ecology, Disposal & Regulatory
| Aquatic Toxicity | Toxic to aquatic organisms with long-lasting effects (H411). Do not discharge to waterways or drains. |
| Disposal | Dispose per local EPA regulations as chemical waste. Adsorb onto activated charcoal before disposal if applicable. |
| Transport | Not classified as dangerous goods — ADG Code, IMDG, IATA |
| Regulatory Framework | Australian WHS Regulations 2023 | GHS 7th Edition | Safe Work Australia COP 2021 |
| AICIS Status | Listed on Australian Inventory of Industrial Chemicals (AIIC) |
| Poisons Schedule | Not scheduled under SUSMP for laboratory use |
Quality Specifications
| Parameter | Specification | Method |
|---|---|---|
| Appearance | Dark purple crystalline powder with metallic green sheen; free-flowing | Visual |
| Dye Content | ≥90% (CI 42555) | Dye content titration |
| λmax | 590 ± 5 nm | UV-Vis spectrophotometry |
| Gram Stain Performance | Clear Gram-positive / Gram-negative differentiation | Functional staining test |
| Solubility | Complete dissolution in water at 10 mg/mL | Visual inspection |
| Heavy Metals | ≤20 ppm | ICP-OES |
| Batch COA | Available for every production lot | On request or download |
Manufacturing & Documentation
Manufactured by AuSaMicS Pty Ltd
31 Longview CT, Thomastown VIC 3074, Australia | ABN 56 676 640 467
✓ Batch-level quality control on every lot
✓ Full GHS-compliant documentation (COA, TDS, SDS)
✓ Australian stock — no import delays, no customs holdups
✓ Technical support direct from our formulation team
31 Longview CT, Thomastown VIC 3074, Australia | ABN 56 676 640 467
✓ Batch-level quality control on every lot
✓ Full GHS-compliant documentation (COA, TDS, SDS)
✓ Australian stock — no import delays, no customs holdups
✓ Technical support direct from our formulation team
Available Documentation
| 📄 Certificate of Analysis (COA) | Batch-specific | Available on request |
| 📋 Technical Data Sheet (TDS) | Full specifications and preparation notes |
| ⚠️ Safety Data Sheet (SDS) | GHS 7th Ed | WHS 2023 compliant | 16 sections |
⚠️ Intended Use: For laboratory and research use only. Not for diagnostic, clinical, therapeutic, human, or veterinary use. Handle with appropriate PPE — CV is a suspected mutagen and carcinogen (Category 2). Avoid skin and eye contact.