Bottom Agar Medium (Ames Test) - BioReagent

Cat. No.: B1528507

GRAU & PUREZA BioReagent, Suitable for microbiology

Synonyms

Bottom Medium | Base Medium | The Underlying Medium(Ames Test)

Storage

Protected from light,Room temperature

Shipped In

Normal

Application

1126

Visão geral

Suitable for the Salmonella‑mammalian microsome enzyme system mutagenicity assay (Ames Test) for genotoxicity detection of test articles.

Medium Formula：

Components	Content
Dextrose	20.0 g/L
Diammonium hydrogen phosphate	3.5 g/L
Dipotassium phosphate	10.0 g/L
Citric acid	2.0 g/L
Magnesium Sulfate	0.2 g/L
Agar	15.0 g/L

Instructions for Use:

Weigh 50.7 g of this product and add into 1 L of distilled or deionized water. Boil for 1 minute to dissolve, make up volume to 1000 mL, dispense into containers, and sterilize at 115 ℃ for 30 minutes. Mix thoroughly immediately after sterilization to prevent agar sedimentation and solidification at the bottom. Set aside for use.

Precautions:

1. This product contains insoluble matter, pH 7.2±0.2 (25 ℃).

2. Avoid direct contact with skin and eyes during handling. In case of accidental contact, immediately flush with plenty of clean water.

3. This product is for laboratory research purposes only and shall not be used for clinical diagnosis.

Especificações

Sinónimos

Bottom Medium | Base Medium | The Underlying Medium(Ames Test)

Especificações e pureza

BioReagent, Suitable for microbiology

Estabilidade e armazenamento

Store at room temperature long term (36 months).Store in the dark.

Condições de armazenamento de armazenamento

Protected from light,Room temperature

Enviado em

Normal

Grau

BioReagent, Suitable for microbiology

Product Line

Produtos biológicos

Propriedades químicas e físicas

• Sensibilidade: Light-sensitive

Application Protocols

The following outlines a general, literature‑based workflow for using Bottom Agar Medium in the Ames test. These are not item‑specific instructions; consult your SOPs and the product’s CoA/Spec Sheet for exact details.

A. Preparation of bottom agar plates (general):

• Reconstitute the medium in high‑purity water; heat to dissolve. Autoclave (121 °C). Cool to ~50–55 °C and pour into sterile Petri dishes to a consistent depth. Allow to solidify and dry. Verify one plate per batch for sterility.

B. Preparation of top agar (separate medium; literature):

• Prepare soft agar (≈0.6–0.7% agar) supplemented with a trace of histidine and biotin. Keep molten at ~45 °C prior to overlay.

C. Assay setup (overlay method; literature):

• Combine: tester strain culture (e.g., Salmonella TA98/TA100/etc.), vehicle or test article (with appropriate solvent control), and ± S9 metabolic activation mix. Add to molten top agar, mix gently, and immediately overlay onto the pre‑poured bottom agar plate.
• Incubate inverted plates at 37 °C (typical) for 48–72 h depending on strain and protocol.
• Include controls: vehicle control, positive controls appropriate to strain and ±S9 (e.g., sodium azide, 2‑nitrofluorene, 2‑aminoanthracene; literature examples).

D. Evaluation:

• Count revertant colonies (manual or automated). Assess cytotoxicity (background lawn thinning) and precipitation. Apply statistical analysis per OECD TG 471 or internal SOP.

Note: Exact plate volumes, incubation times, and control concentrations are protocol‑dependent and not specified for this item.

Biological Roles

This product functions as a minimal growth substrate for bacterial tester strains used in the Ames assay. It does not have a single defined biological role at the molecular level because it is a mixture, but its components collectively influence microbial physiology and assay selectivity.

General biological functions (literature; composition may vary by supplier):

• Carbon source (often glucose): Supports limited growth sufficient for DNA replication and expression of revertant phenotypes without allowing confluent lawn formation.
• Inorganic salts and phosphate: Provide essential ions and buffering capacity to maintain physiological pH during incubation.
• Magnesium and trace ions: Support enzymatic functions (e.g., DNA repair, replication) important for detecting mutagenic events.
• Agar matrix: An inert, non‑nutritive gel that creates a stable two‑dimensional surface for colony isolation and accurate counting.

Assay‑specific considerations:

• Histidine omission in the bottom agar enforces selection; only top agar supplies trace histidine/biotin (overlay method), ensuring revertant colonies derive from true back‑mutations or frameshift reversion.
• Ionic strength and pH closer to neutrality favor Salmonella viability while maintaining selection pressure; exact targets depend on the validated recipe.

Limitations/controls:

• Background spontaneous reversion varies by strain; standardized bottom agar helps maintain consistent baseline.
• Vehicle controls and positive controls (e.g., sodium azide, 2‑nitrofluorene, 9‑aminoacridine; literature examples) are essential to interpret results independent of medium batch effects.

Buffer Applications

This product is a culture medium rather than a dedicated buffer. While minimal media commonly include phosphate salts that provide some buffering capacity near neutrality (literature), Bottom Agar Medium is not typically used as a standalone buffer solution.

• If a buffered aqueous phase is needed for sample preparation or dosing solutions in the Ames test, prepare separate, well‑defined buffers (e.g., phosphate buffer at target pH) according to validated SOPs.
• Refer to the item’s CoA/Spec Sheet for any stated pH of the reconstituted medium; item‑specific buffer capacity is not specified for this product.

Green Alternatives

While culture media are not typically framed in terms of green solvent substitution, several sustainability considerations apply to Bottom Agar Medium usage.

• Renewable biopolymer: Agar is derived from red seaweeds (renewable biomass) and is biodegradable. This compares favorably to synthetic gelling agents derived from petrochemicals.
• Ready‑to‑use vs. in‑house prep:
  • Pre‑poured plates reduce energy use from repeated autoclaving but increase packaging and cold‑chain needs (if supplied sterile). In‑house pouring allows batching and container reuse but requires autoclave energy/water.
• Process efficiency:
  • Optimize batch size to minimize waste from expired plates.
  • Use reusable glassware for media prep and dispensing to reduce single‑use plastics, balancing with sterilization water/energy cost.
• Decontamination and waste:
  • Consolidate autoclave loads; verify spore‑test performance to avoid re‑runs.
  • Segregate uncontaminated packaging for recycling when permitted.

Comparison (general, not item‑specific):

• Agar vs. gellan gum: Gellan can form clear gels at lower concentration, potentially reducing material and energy to melt; supply‑chain impacts vary. Agar remains widely available and robust across ionic strengths.
• Dry powder media vs. liquid: Shipping dry media avoids weight/volume and reduces transportation emissions; reconstitute on site.

Environmental note: Implement local green‑lab practices (e.g., LEAF/My Green Lab) to improve overall sustainability independent of the specific medium chosen.

Pharmaceutical Uses

No therapeutic or clinical use is intended. For research use only (per Product Data).

Relevant non‑clinical/pharmaceutics context (general):

• Mutagenicity screening: Bottom agar plates are integral to the Ames test (OECD TG 471), widely used in pharmaceutical R&D and quality/safety assessment to screen drug candidates, impurities, process intermediates, and excipients for mutagenic potential.
• Manufacturing support: Can be used in development laboratories to evaluate extractables/leachables or degradation products using bacterial reverse‑mutation assays.

Regulatory/quality notes:

• While the Ames test is part of ICH S2(R1) guidance for genotoxicity testing strategies, this specific product is not certified for regulated testing unless your quality system qualifies it. Establish vendor qualification, retain CoA, and perform incoming QC as required by your QA procedures.
• Documentation: Record medium lot numbers on all assay records to enable traceability in investigations and trend analyses.

Excipient role: None. This medium is not an excipient or formulation additive.

Any pharmacopeial monographs do not apply to this composite medium. For detailed specifications and suitability in a regulated workflow, consult the CoA/Spec Sheet and validate per your internal protocols.

Physical Properties

Item‑specific physical constants are not provided in the Product Data for this complex medium.

• Appearance: Not specified for this item; refer to CoA/Spec Sheet.
• Solubility: Water‑reconstitutable (general for agar media; components dissolve/disperse in hot water). Item‑specific behavior: Not specified for this item.
• pH (reconstituted): Not specified for this item; refer to CoA/Spec Sheet.
• Density, BP, MP, refractive index, UV cutoff: Not applicable/not specified for this mixture; refer to CoA/Spec Sheet.

Typical behavior and benchmarks (literature/general; NOT specifications):

• Agar gelling: Solutions generally gel on cooling; literature gel point ~32–40 °C and melt point ~80–90 °C for standard agar, depending on concentration and agaroid composition.
• Working state: Media are typically brought to a rolling boil or autoclaved (121 °C) to fully dissolve/sterilize, then poured at ~50–55 °C to form plates with smooth surfaces.
• Osmolarity/ionic strength: Determined by the minimal salts recipe; exact values vary with formulation.
• Color: Minimal agar media are often off‑white to pale straw as powders and form translucent, slightly opalescent gels when poured (literature).

Notes for users:

• Use high‑purity water to minimize precipitation or haze.
• Avoid overheating/evaporation that can shift salt concentrations and pH. Always calibrate to your lab’s validated method or the item’s CoA/specification sheet.

Quality and Grades

• Item grade: BioReagent (per Product Data).

What BioReagent grade generally implies (general guidance):

• Sourced and processed to meet typical requirements for biological and microbiological research. Emphasis on lot‑to‑lot consistency, low bioburden prior to sterilization, and performance in defined assays or growth tests.
• Suitable for preparing culture plates used in the Ames mutagenicity assay, where background colony counts and surface quality are critical metrics.

How to qualify this item in your lab:

• Verify solidification and plate quality: clarity, absence of crystals/precipitates, uniform surface.
• Performance checks (literature guidance): minimal background growth of tester strains without histidine/biotin overlay; consistent revertant recovery when used with standardized top agar and controls.
• Documentation: For exact acceptance criteria (e.g., moisture content, pH after reconstitution, growth performance), consult the product’s CoA/Spec Sheet. If parameters are missing, establish in‑house QC assays (e.g., sterility checks, pour reproducibility, plate hardness tests).

Stabilizers/additives:

• Not specified for this item; refer to CoA/Spec Sheet for any antimicrobial inhibitors, anti‑caking agents, or pH adjusters.

Regulatory note:

• For research use only (per Product Data). Not intended for diagnostic or clinical use. If you operate under GLP/GMP, qualify and document the medium per your quality system.

Reaction and Applications

Chemical reaction use is not applicable. This product is intended as the solid base (“bottom agar”) for the Ames bacterial reverse‑mutation assay.

Ames test application (literature/general):

• Purpose: Detects mutagenic potential of chemicals by measuring reversion of histidine‑auxotrophic Salmonella typhimurium (and sometimes E. coli WP2 uvrA) to prototrophy.
• Role of bottom agar: Provides a minimal, histidine‑free solid substrate that limits background lawn growth. Revertant colonies arise on the surface after overlaying with top agar containing a trace amount of histidine/biotin and the test article (± metabolic activation with S9).
• Why minimal: Suppresses non‑revertant growth and accentuates selection pressure, improving signal‑to‑noise for revertant scoring.
• Practical tips (general, not item‑specific):
  • Ensure plates are dry (surface free of condensation) before use; moisture can cause colony spreading and merging.
  • Pour uniform plate thickness to standardize diffusion and colony size.
  • Label and segregate plates by strain and S9 condition; avoid cross‑contamination.
  • Validate plate sterility by incubating one uninoculated plate per batch.

Compatibility:

• Designed to be used with standard Ames top agar overlays (histidine/biotin supplement, 0.6–0.7% soft agar; literature) and with metabolic activation systems (e.g., Aroclor‑induced rat liver S9) per OECD TG 471.

Note: For test‑article‑specific solubility/vehicle effects, consult relevant guidance (OECD 471) and your lab’s SOPs.

Reaction Conditions

Chemical reaction conditions are not applicable. The relevant “conditions” here pertain to media preparation and plate pouring for biological assays (general literature guidance; NOT item‑specific specifications):

• Reconstitution: Add dry medium to high‑purity water with stirring. Heat to dissolve completely. Exact grams per liter and target pH are not specified for this item; refer to CoA/Spec Sheet and your SOP.
• Sterilization: Autoclave at 121 °C for an appropriate holding time (commonly 15–20 minutes for volumes ≤1 L; literature). Avoid over‑sterilization that can cause Maillard browning or caramelization of sugars.
• Pouring temperature: Cool molten agar to ~50–55 °C before pouring plates to minimize condensation and preserve agar strength.
• Plate thickness: Maintain a consistent volume per plate to standardize diffusion and colony morphology across experiments.
• Drying/conditioning: Allow plates to set and dry (e.g., overnight at room temperature in sterile conditions) or briefly in a laminar‑flow hood before use to prevent surface pooling of the overlay top agar.
• Storage of poured plates: Store inverted, protected from light and desiccation, at 2–8 °C or room temperature per your validated practice; exact storage for this item is not specified—consult CoA/Spec Sheet. Use within your validated shelf‑life to limit dehydration and pH drift.

Controls: Prepare at least one uninoculated plate per batch to check sterility and surface quality before assay setup.

Safety and Handling

GHS classification, signal word, pictograms, and H‑statements are not provided in the Product Data. Always consult the item‑specific SDS for authoritative safety information.

General laboratory safety guidance for dry culture media (not item‑specific):

• Hazards: Nuisance dust may cause mechanical irritation to eyes/respiratory tract. Spilled dry powder can be slippery when wet. Non‑sterile medium can support microbial growth after reconstitution.
• PPE: Lab coat, safety glasses, and suitable gloves (e.g., nitrile). Use a dust mask/respirator if handling large quantities that may aerosolize (per risk assessment).
• Handling: Avoid creating dust. Reconstitute with water in heat‑resistant glassware; use appropriate thermal protection when handling hot molten agar. Mix gently to minimize foaming.
• Sterilization and hygiene: Autoclave as required by your protocol (typical 121 °C), and maintain aseptic technique to prevent contamination. Disinfect work surfaces after handling.
• Incompatibilities: No specific chemical incompatibilities are typical for agar media; avoid contact with strong oxidizers or acids/bases that could alter pH and gel properties (general precaution).
• First aid (general):
  • Eye contact: Rinse cautiously with water for several minutes; remove contact lenses if present and easy to do; seek medical attention if irritation persists.
  • Skin contact: Wash with soap and water; for thermal burns from hot agar, cool with running water and seek medical attention.
  • Inhalation: Move to fresh air; seek medical advice if symptoms occur.
  • Ingestion: Rinse mouth; obtain medical attention if unwell.
• Waste: Autoclave biologically contaminated media/plates prior to disposal per institutional biosafety procedures.

Solvent Selection

This product is a dehydrated culture medium designed to be reconstituted with water; selection of organic solvents is not applicable.

Water quality considerations (applicable):

• Use freshly produced Type I or equivalent high‑purity water (low TOC, low ionic contaminants) to prevent precipitation and to maintain reproducibility of ionic strength.
• Avoid chelators or residual detergents from glassware that can interfere with divalent cations (e.g., Mg2+) important for bacterial physiology and plating efficiency.
• Degassing is optional; gentle warming facilitates dissolution and minimizes bubble formation in poured plates.

Practical solvent/use scenarios:

• Reconstitution: Add the medium to cold or room‑temperature water with stirring; heat to dissolve fully before sterilization.
• Sterilization: Autoclave in appropriate vessels; do not use organic solvents.
• Pouring: Cool molten medium to ~50–55 °C before pouring to reduce condensation and maintain agar integrity (literature practice).

Comparison to alternatives (literature/general):

• Minimal bottom agar vs. rich media (e.g., LB, TSA): Minimal agar reduces background growth in Ames assays and provides selective pressure for revertants; rich media increase background and are not recommended for the base layer.

For precise reconstitution volumes and conditions: Not specified for this item; refer to CoA/Spec Sheet and your validated SOP.

Storage and Reconstitution

• Storage conditions (as sold): Room temperature (per Product Data). Protect from moisture and excessive heat. Keep container tightly closed to prevent caking and contamination.
• Shipping: Normal (per Product Data).
• Research use: For research use only (per Product Data).

Reconstitution (general guidance; NOT item‑specific specifications):

• Water: Use freshly prepared Type I water. Rinse vessels to remove detergents/chelators.
• Dissolution: Add the powder to measured water with stirring; heat to fully dissolve. Avoid scorching. If necessary, adjust pH per your validated recipe after dissolution (exact targets are not specified for this item; refer to CoA/Spec Sheet/SOP).
• Sterilization: Autoclave at 121 °C using validated time/volume settings. Mix gently post‑autoclave to re‑homogenize before pouring.
• Pouring: Cool to ~50–55 °C and dispense into sterile Petri dishes. Allow to solidify and dry to the desired surface moisture.

Post‑pour storage (general):

• Store plates inverted in sealed sleeves to limit dehydration and contamination. Temperature and shelf‑life are method‑dependent and not specified for this item; refer to CoA/Spec Sheet and your lab’s validated practice.

Not specified for this item; refer to CoA/Spec Sheet for:

• Exact reconstitution concentration (g/L),
• Target pH after preparation,
• Appearance and performance criteria,
• Shelf‑life of powder and poured plates.

Structure and Identity

Bottom Agar Medium (Ames Test) is a formulated culture medium used as the solid base layer in the Salmonella reverse‑mutation (Ames) assay. It is a complex mixture rather than a single chemical entity.

• SKU: B1528507
• Product name: Bottom Agar Medium (Ames Test)
• Category: Life Science reagents; microbiology/media component for mutagenicity testing
• Grade/Purity: BioReagent (per Product Data)
• Molecular formula: Not applicable (mixture)
• Molecular weight: Not applicable (mixture)
• CAS, CID, InChIKey, SMILES: Not specified for this item; refer to CoA/Spec Sheet.

Structural/functional description (general knowledge):

• The medium typically comprises a minimal salts base solidified with agar. In the Ames test, this “bottom agar” provides a nutrient‑limited surface that discourages lawn growth of histidine‑auxotrophic Salmonella strains and allows detection of revertant colonies.
• Key functional components (literature, may vary by supplier):
  • Minimal inorganic salts (e.g., phosphate buffer, ammonium salts, Mg2+),
  • A defined carbon source (commonly glucose),
  • Agar as the gelling agent.

2D/functional depiction in words (general):

• Heterogeneous granular/powdered medium whose dissolved ions and small molecules create a minimal environment; the agar polysaccharide forms a 3D gel matrix upon cooling from molten solution. No stereochemical or ring‑system description is applicable to the overall mixture.

Synthetic Utility

Not applicable. Bottom Agar Medium (Ames Test) is not a chemical building block or reagent for synthetic transformations. It serves as a biological culture matrix in microbiology/genotoxicity assays.

For users seeking synthetic chemistry content:

• Consider referring to the Reaction & Applications and Application Protocols sections for assay‑related usage.
• If your objective is to study chemical reactivity of a test article, the Ames assay uses this medium as an inert platform; all reactions of interest occur within the cells or through metabolic activation (e.g., S9), not in the medium itself.

Target Specificity

Not applicable. This product is not a biochemical probe, enzyme inhibitor, or antibody and has no molecular target specificity. It is a general culture substrate for bacterial tester strains used in the Ames assay.