TBX Agar (AS-1427) | Chromogenic E. coli Enumeration
TBX Agar (Tryptone Bile X-glucuronide Agar)
Chromogenic Medium for Rapid Detection of Escherichia coli (ISO 16649)
Catalog Number: AS-1427 | HS Code: 3821.00.00 | Standard: ISO 16649 | Format: Dehydrated Powder
Overview
TBX Agar (Tryptone Bile X-glucuronide Agar) is a selective chromogenic culture medium designed for the rapid detection and enumeration of Escherichia coli in water, food, and environmental samples. The medium exploits the enzymatic activity of β-glucuronidase (GUD), which is produced by the vast majority of E. coli strains, to generate a distinct blue-green coloration upon hydrolysis of the chromogenic substrate X-glucuronide. TBX Agar is compliant with ISO 16649 for the enumeration of E. coli in food and animal feeding stuffs.
Principle of the Medium
TBX Agar contains tryptone as a nitrogen and nutrient source, and bile salts to inhibit the growth of Gram-positive organisms. The selective action of bile salts ensures that only Gram-negative bacteria, primarily coliforms and E. coli, are recovered.
The key functional component is 5-bromo-4-chloro-3-indolyl-β-D-glucuronide (X-glucuronide), a chromogenic substrate cleaved specifically by β-glucuronidase. Upon enzymatic hydrolysis, an insoluble indigo-blue chromophore precipitates at the colony site, producing the characteristic blue-green coloration that allows direct visual identification of E. coli without the need for further confirmatory tests.
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Insoluble indigo chromophore released
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Blue-green colonies = E. coli confirmed
Colony Appearance
| Organism | Colony Appearance | Interpretation |
|---|---|---|
| Escherichia coli | 🔵 Blue-green | GUD-positive — confirmed E. coli |
| Other coliforms (Klebsiella, Enterobacter) | ⚪ Colourless / pale | GUD-negative — not E. coli |
| Gram-positive organisms | ❌ Inhibited | Suppressed by bile salts |
| E. coli O157:H7 | ⚪ Colourless or pale | GUD-negative variant — may not be detected |
Key Features & Benefits
Rapid Detection
- Direct visual confirmation — no subculture needed
- Results typically within 18–24 hours at 44°C
- Reduces time-to-result vs. conventional media
Selectivity & Specificity
- Bile salts inhibit Gram-positive background flora
- Chromogenic substrate is highly specific for GUD
- Clear discrimination between E. coli and other coliforms
Regulatory Compliance
- ISO 16649 compliant methodology
- Suitable for accredited food and water testing workflows
- Full documentation set (COA, TDS, SDS) provided
Versatility
- Compatible with membrane filtration (MF) and pour plate methods
- Applicable to water, food, dairy, meat, and environmental samples
- Available as dehydrated powder — flexible batch preparation
Typical Applications
- Detection and enumeration of E. coli in drinking water, wastewater, and recreational water
- Food safety testing — dairy, meat, beverages, produce
- Environmental monitoring of surfaces and air in food production facilities
- Membrane filtration and pour plate enumeration workflows
- Quality control (QC) laboratories in food manufacturing and water utilities
- Research and teaching in applied and environmental microbiology
Preparation Guidelines
| Parameter | Recommended Condition |
|---|---|
| Dissolution | Suspend powder in distilled water; heat to boiling with stirring until fully dissolved |
| Sterilisation | Autoclave at 121°C for 15 minutes |
| Pour temperature | Cool to 50–55°C before pouring |
| Incubation temperature | 44 ± 0.5°C |
| Incubation time | 18–24 hours |
| Atmosphere | Aerobic |
Technical Overview & Biochemistry
β-Glucuronidase (GUD, EC 3.2.1.31) is a lysosomal enzyme that cleaves the β-glycosidic bond of β-D-glucuronic acid conjugates. Approximately 94–97% of E. coli strains are GUD-positive, making GUD activity a highly reliable marker for this species. The uidA gene encodes GUD in E. coli, and its activity is constitutively expressed under the conditions used in TBX Agar. Bile salts (deoxycholate and cholate mixtures) act by disrupting the cytoplasmic membrane of Gram-positive bacteria while leaving Gram-negative organisms largely unaffected, providing a robust selective environment.
E. coli Detection Media Comparison
| Medium | Detection Principle | Speed | Best Use | Manufacturer Equivalent |
|---|---|---|---|---|
| TBX Agar (AS-1427) | Chromogenic (GUD / β-glucuronidase) | Fast (18–24 h) | ISO 16649 E. coli detection | AuSaMicS, Melbourne |
| MacConkey Agar | Lactose fermentation / bile salts | Moderate | General coliform recovery | Oxoid CM0007, BD 212123 |
| EMB Agar | Dye precipitation (eosin/methylene blue) | Moderate | Coliform differentiation / metallic sheen | Oxoid CM0069, BD 221065 |
| Chromogenic Coliform Agar | Multiple chromogenic substrates | Very fast | Simultaneous coliforms + E. coli | Oxoid PO5038, Merck 1.10426 |
| mFC Agar | Faecal coliform growth at 44.5°C | Moderate | Water membrane filtration (APHA) | Oxoid CM0665, BD 214757 |
Cross-Reference / Equivalent Products
| Brand | Equivalent Product | Catalogue Number |
|---|---|---|
| Oxoid (Thermo Fisher) | TBX Agar | CM0945 |
| BD Difco | TBX Agar | — |
| Merck | TBX Agar | 1.05267 |
| Thermo Fisher | Chromogenic E. coli / Coliform Agar | PO5038A |
Storage & Shelf Life
- Store at 15–25°C in a dry location, away from heat and moisture
- Keep container tightly sealed when not in use
- Protect from direct light exposure
- Shelf life: stated on label; refer to COA for batch-specific expiry
Intended Use
For laboratory and research use only. Not for direct human, veterinary, or food additive use.
Marketing / Display Titles
- TBX Agar (AS-1427) – ISO 16649 Chromogenic Medium for E. coli Detection | AuSaMicS Australia
- Tryptone Bile X-glucuronide Agar – Rapid E. coli Confirmation in Water & Food Testing
- Australian-Made TBX Agar | β-Glucuronidase Chromogenic Detection Medium
Intent-Based Long-Tail Search Queries
- TBX agar Australia supplier
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Technical Data Sheet (TDS) — AS-1427
The Technical Data Sheet for TBX Agar (AS-1427) covers physicochemical composition, quality control performance data, mode of action, and peer-reviewed literature references in accordance with AuSaMicS TDS Option 2 documentation standards.
Composition (per litre of prepared medium)
| Component | Function | Approximate Amount |
|---|---|---|
| Tryptone | Nitrogen and growth nutrient source | 20.0 g |
| Bile salts (mixed) | Selective inhibitor of Gram-positive organisms | 1.5 g |
| X-glucuronide (5-bromo-4-chloro-3-indolyl-β-D-glucuronide) | Chromogenic substrate for β-glucuronidase | 0.075 g |
| Agar | Solidifying agent | 15.0 g |
| Final pH | — | 7.2 ± 0.2 at 25°C |
Physicochemical Specifications
| Parameter | Specification |
|---|---|
| Appearance (powder) | Beige to cream-coloured, free-flowing powder |
| Appearance (prepared medium) | Light amber, clear to slightly opalescent gel |
| pH (prepared, 25°C) | 7.2 ± 0.2 |
| Gelling | Firm gel at incubation temperature (44°C) |
| Moisture content (powder) | ≤ 6% |
| Storage temperature | 15–25°C, dry |
Quality Control Performance
| Test Organism | ATCC / Reference Strain | Inoculum (CFU) | Expected Result |
|---|---|---|---|
| Escherichia coli | ATCC 25922 | ≤ 100 | Blue-green colonies; good growth |
| Escherichia coli O157:H7 | ATCC 35150 | ≤ 100 | Colourless colonies (GUD-negative) |
| Klebsiella pneumoniae | ATCC 13883 | ≤ 100 | Colourless colonies; partial inhibition |
| Enterococcus faecalis | ATCC 29212 | ≤ 100 | Inhibited; no growth |
| Staphylococcus aureus | ATCC 25923 | ≤ 100 | Inhibited; no growth |
Mode of Action
TBX Agar operates through two complementary mechanisms. First, bile salts suppress the growth of Gram-positive organisms by disrupting their cytoplasmic membranes, providing selective enrichment of Gram-negative enteric bacteria. Second, the chromogenic substrate X-glucuronide (5-bromo-4-chloro-3-indolyl-β-D-glucuronide) is hydrolysed by β-glucuronidase (encoded by the uidA gene), an enzyme constitutively expressed by approximately 94–97% of E. coli strains at 44°C. The released indoxyl moiety spontaneously dimerises and oxidises to form an insoluble blue-green dye (5,5'-dibromo-4,4'-dichloro-indigo), precipitating at the colony site and providing direct visual confirmation of E. coli.
Literature References
- Manafi, M., & Ossmer, R. (1993). Fluorocult TBX Agar: a new chromogenic medium for the rapid detection of E. coli in water. Zentralblatt für Hygiene und Umweltmedizin, 194(4), 380–386.
- ISO 16649-2:2001. Microbiology of food and animal feeding stuffs — Horizontal method for the enumeration of β-glucuronidase-positive Escherichia coli. International Organisation for Standardisation, Geneva.
- Frampton, E.W., & Restaino, L. (1993). Methods for E. coli identification in food, water and clinical samples based on beta-glucuronidase detection. Journal of Applied Bacteriology, 74(3), 223–233.
Full TDS document (five-file documentation set: PD, TDS, SDS, COA + HTML embed) available as a downloadable PDF.
Safety Data Sheet (SDS) — Summary
The full Safety Data Sheet for TBX Agar (AS-1427) is prepared in accordance with the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) and Australian WHS Regulations (Safe Work Australia), covering all 16 mandatory sections.
| Section | Summary |
|---|---|
| 1. Identification | TBX Agar (Tryptone Bile X-glucuronide Agar), AS-1427. AuSaMicS Pty Ltd, 31 Longview CT, Thomastown VIC 3074. support@ausamics.com | +61 412 520 598 |
| 2. Hazard Identification | Not classified as a hazardous substance under Australian WHS legislation. No GHS hazard pictograms apply. Signal Word: Not required. |
| 3. Composition | Mixture: tryptone, bile salts, X-glucuronide, agar. No listed hazardous substances above threshold limits. |
| 4. First Aid Measures | Inhalation: move to fresh air. Skin/eye contact: wash with water. Ingestion: seek medical advice. No specific antidote required. |
| 5. Fire-Fighting Measures | Not flammable. Use appropriate extinguishing media for surrounding fire. |
| 6. Accidental Release | Powder: avoid dust generation; sweep/vacuum and place in sealed container. Avoid release to waterways. |
| 7. Handling & Storage | Store at 15–25°C in a dry location; keep container tightly sealed. Avoid inhalation of powder dust. Use in a well-ventilated laboratory environment. |
| 8. Exposure Controls / PPE | Standard laboratory PPE: nitrile gloves, safety spectacles, laboratory coat. Local exhaust ventilation recommended when handling powder in quantity. |
| 9. Physical & Chemical Properties | Form: powder. Colour: beige to cream. Odour: faint. pH (prepared): 7.2 ± 0.2. Solubility: dispersible in water with heat. |
| 10. Stability & Reactivity | Stable under recommended storage conditions. Avoid strong oxidising agents and excessive moisture. |
| 11. Toxicological Information | No acute or chronic toxicity data indicating significant hazard at normal laboratory use quantities. |
| 12. Ecological Information | Not expected to be harmful to the environment at typical use levels. Avoid large-scale release to waterways. |
| 13. Disposal Considerations | Dispose of in accordance with local council and state EPA regulations. Prepared agar containing microbiological cultures must be autoclaved prior to disposal. |
| 14. Transport Information | Not classified as a dangerous good under ADG/IATA/IMDG regulations. |
| 15. Regulatory Information | For laboratory and research use only. Compliant with Australian WHS Regulations and GHS (7th revised edition). |
| 16. Other Information | Prepared by AuSaMicS Pty Ltd. Refer to full SDS document for complete regulatory and safety information. |
Full 16-section GHS-compliant SDS with embedded GHS pictograms available as a downloadable PDF.
Certificate of Analysis (COA) — AS-1427
Each batch of TBX Agar (AS-1427) is produced under quality-controlled conditions and is released with a batch-specific Certificate of Analysis confirming compliance with AuSaMicS physicochemical and microbiological performance specifications.
COA — Typical Specifications
| Test Parameter | Specification | Method | Result |
|---|---|---|---|
| Appearance (powder) | Beige to cream, free-flowing | Visual | Conforms |
| pH (prepared, 25°C) | 7.2 ± 0.2 | Potentiometric | Within specification |
| Moisture content | ≤ 6% | Loss on drying | Within specification |
| Gel consistency | Firm gel at 44°C incubation | Visual observation | Conforms |
| E. coli ATCC 25922 (GUD+) | Blue-green colonies; ≥ 70% recovery (≤ 100 CFU inoculum) | Pour plate, 44°C, 18–24 h | Conforms |
| E. coli O157:H7 ATCC 35150 | Colourless colonies | Pour plate, 44°C, 18–24 h | Conforms |
| Enterococcus faecalis ATCC 29212 | Inhibited; no growth | Pour plate, 44°C, 18–24 h | Conforms |
| Staphylococcus aureus ATCC 25923 | Inhibited; no growth | Pour plate, 44°C, 18–24 h | Conforms |
Batch Traceability
| Field | Detail |
|---|---|
| Product | TBX Agar (Tryptone Bile X-glucuronide Agar) |
| Catalogue Number | AS-1427 |
| Manufacturer | AuSaMicS Pty Ltd, 31 Longview CT, Thomastown VIC 3074, Australia |
| Batch Number | Stated on label |
| Manufacturing Date | Stated on label |
| Expiry Date | Stated on label |
| Storage Conditions | 15–25°C, dry, away from light and moisture |
| Intended Use | For laboratory and research use only |
Batch-specific COA is issued at the time of dispatch and is available for download or provided as a physical document on request.
AuSaMicS Pty Ltd | 31 Longview CT, Thomastown, VIC 3074, Australia | ABN: 56 676 640 467 | www.ausamics.com.au | support@ausamics.com | +61 412 520 598
All products manufactured under quality-controlled conditions. Specifications subject to change without notice. For laboratory and research use only. AuSaMicS Pty Ltd accepts no liability for misuse of this product outside its intended laboratory application.