Creatine Content Assay Kit (Micro Method)

Cat. No.: C1522092

グレード＆純度 BioReagent

概要

Creatine is a nitrogen-containing organic acid primarily stored in the muscles and brains of vertebrate animals, where it participates in energy metabolism (by rapidly generating ATP through phosphocreatine). Creatine levels in animals are influenced by diet, exercise, species, age, and health status. This product provides a simple, convenient, and rapid method for detecting creatine content, suitable for animal tissue and serum (plasma) samples. Its principle is based on the reaction of creatine with diacetyl-α-naphthol under alkaline conditions, producing a red product with an absorption peak at 520 nm.

仕様

仕様と純度

BioReagent

等級

BioReagent

アプリケーション

Cell Metabolism

サンプルタイプ

プラズマ, 動物組織, 血清

検出方法

Colorimetric Method

Detection instrument

Microplate reader,Visible Spectrophotometer

Detection wavelength

520nm

保管と配送

保管条件

Store at 2-8°C,Protected from light

入荷

Wet ice

安定性と保管

Store at 2-8℃ for 6 months. Store in the dark.

Application Protocols

Item-specific tested applications/dilutions: Not specified for this item; refer to the kit insert.

General example protocol (literature; for planning only—follow the kit insert for exact steps)

1. Preparation

• Equilibrate reagents to room temperature (keep enzymes on ice during setup). Protect dye solutions from light.
• Prepare a creatine standard curve (e.g., serial dilutions covering expected sample range) in assay buffer.

2. Sample handling

• Clarify samples by centrifugation. If needed, deproteinize (PCA/TCA or ultrafiltration) and neutralize. Avoid azide and strong reductants.

3. Plate setup (96-well)

• Allocate wells for Blank, Standards (≥5 levels), and Samples (≥2 replicates each).
• Add sample or standard (e.g., 10–50% of final well volume), then add working reagent mix to reach 50–200 µL per well.

4. Incubation

• Mix gently and incubate at RT or 37°C for 10–40 min until color develops. Protect from light.

5. Measurement

• Read absorbance at the recommended wavelength (commonly 540–600 nm depending on dye system).

6. Calculations

• Subtract Blank absorbance; construct the standard curve; interpolate sample concentrations. Apply dilution factors. Report in µM, nmol/well, or mg/g depending on matrix.

Quality controls

• Include spike-recovery and inter-plate controls to monitor accuracy and precision over time.

Biological Roles

General biology of the analyte (creatine) — literature overview (not kit-specific)

• Function: Creatine serves as a high-energy phosphate buffer via the phosphocreatine/creatine kinase system, facilitating rapid ATP regeneration in tissues with fluctuating energy demand (muscle, brain).
• Metabolism: Synthesized primarily from arginine, glycine, and methionine; transported to target tissues and reversibly phosphorylated by creatine kinases. Non-enzymatic cyclization yields creatinine at a relatively constant rate.
• Compartmentation: Cytosolic and mitochondrial creatine kinase isoforms support intracellular energy shuttling; creatine/phosphocreatine helps maintain ATP/ADP ratios and buffer energy transients.
• Homeostasis and transport: Creatine transporter (SLC6A8) mediates uptake; dietary intake and endogenous synthesis both contribute to pools.

Experimental implications

• Sample handling can influence measured creatine due to enzymatic/chemical interconversion with phosphocreatine and creatinine. Rapid quenching or deproteinization may be required to “freeze” metabolite states for accurate quantitation.
• Matrix interferences (proteins, hemoglobin, colored compounds) may affect colorimetric readouts; include appropriate blanks and recovery controls.

Note: This section provides biological context for the target analyte of the assay kit and is not a statement of clinical use or diagnostic performance.

Buffer Applications

Applicability

• The product is an assay kit; it is not a general buffering agent. However, buffer choice can materially impact assay performance.

General buffer guidance for creatine colorimetric/enzymatic assays (literature)

• pH: Many creatine-oxidase/creatinase systems operate optimally near neutral pH (≈7.0–7.8). Verify the recommended pH in the kit insert; do not substitute buffers unless validated.
• Composition: Avoid sodium azide in any buffer when peroxidase-coupled detection is used, as azide inhibits HRP and quenches signal. Limit strong chelators or high phosphate if metal-dependent enzymes are involved (chemistry dependent).
• Ionic strength: Maintain consistent ionic strength between standards and samples to minimize matrix effects.
• Sample cleanup: For protein-rich matrices, deproteinize via acid precipitation (PCA/TCA) with subsequent neutralization, or employ centrifugal ultrafiltration; ensure the final buffer conditions match the assay working buffer prior to measurement.

Item-specific buffer components, if any, are Not specified for this item; refer to CoA/Spec Sheet.

Green Alternatives

Context

• As a “micro method,” this kit inherently reduces reagent volumes and waste relative to macro or cuvette-based assays, aligning with green chemistry principles of waste minimization.

Greener practices for colorimetric metabolite assays (general guidance)

• Volume reduction: Use 96- or 384-well microplates to minimize solvent/reagent consumption and waste streams.
• Aqueous systems: Prefer water-based buffers over organic-rich formulations; most creatine assays are fully aqueous.
• Hazard reduction: Avoid heavy-metal-based color reagents and limit use of corrosive deproteinization agents; consider ultrafiltration as a cleanup alternative when compatible.
• Energy efficiency: Conduct incubations at ambient temperature if validated for your samples (some assays require 37°C for kinetics; follow the kit insert).

Balanced considerations

• While microplate assays reduce waste, sensitivity and signal-to-noise can be affected by shorter pathlengths. Optimize plate type (clear, flat-bottom; low-binding plastics) and reading mode to maintain performance without increasing reagent consumption.

Note: No kit-specific substitutions are recommended here; follow the provided instructions. The above are general greener-practice guidelines.

Pharmaceutical Uses

Not typically applicable as a drug or excipient. This is a research-use assay kit.

Relevant non-clinical/QA contexts (general information)

• Raw material QC: Colorimetric creatine assays are commonly used to assess creatine content in nutritional raw materials (e.g., creatine monohydrate) as part of supplier qualification or in-process checks.
• Process monitoring: Bioprocess development may track creatine/creatinine pools in cell culture media to understand nitrogen balance or metabolic state (research use only).
• Method transfer: Microplate-based colorimetric methods can be adapted for high-throughput screening and automated liquid handling in R&D quality assessments.

Note: No pharmacopeial status, validated method parameters, or regulatory claims are provided for this kit. It is for research use only and not for use in diagnostic procedures or GMP release testing unless independently validated by the end-user.

Physical Properties

Item-specific facts (from Product Data)

• Physical state/appearance of individual kit components: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular formula and molecular weight of the kit: Not applicable (multi-component kit).

General/literature values (for the analyte or typical assay readouts; not item-specific specifications)

• Creatine (analyte) MW: ~131.13 g/mol; monohydrate MW: ~149.15 g/mol (literature).
• Solubility (creatine): water soluble; increased solubility when warmed or adjusted to neutral pH (literature).
• Typical colorimetric micro-assay readout: absorbance in the visible range (commonly ~540–600 nm depending on dye system) or fluorescence (Ex/Em varies by dye) (literature).
• Typical pathlengths: 96-well plates (200–300 µL per well) or low-volume microplates (≤100 µL per well) (literature).

Method performance considerations (literature; not item-specific)

• Linearity is commonly established with multiple creatine standards spanning low µM to high µM or low mM, depending on sensitivity.
• Detection limit and dynamic range are assay-chemistry dependent; Not specified for this item; refer to CoA/Spec Sheet.

Note: In the absence of a kit-specific specification sheet, do not treat any of the above as certified specifications for SKU C1522092.

Quality and Grades

Item-specific facts (from Product Data)

• Grade: BioReagent.

What BioReagent grade generally implies (general information)

• BioReagent-grade materials are suitable for biological research workflows, emphasizing low levels of impurities that could inhibit enzymes or interfere with spectrophotometric detection. Typical expectations include:
  • Consistency between lots for assay response.
  • Reduced background absorbance/fluorescence in the intended detection window.
  • Compatibility with enzymatic reactions (if enzymes are included).

Kit-specific quality (not specified for this item)

• Exact acceptance criteria (e.g., standard curve linearity, lower limit of detection, inter-assay CV) are Not specified for this item; refer to CoA/Spec Sheet.
• Stabilizers or preservatives (e.g., BSA, glycerol, thimerosal/ProClin/bronopol) may be present in enzyme or dye solutions in typical kits; composition is Not specified for this item.

Recommendations

• Verify lot-specific performance with a standard curve on receipt.
• Record plate reader settings, incubation times, and temperatures to ensure reproducibility across runs and lots.

Reaction and Applications

Item-specific facts (from Product Data)

• Intended use: Quantitative measurement of creatine content by a micro (low-volume) method; For research use only.

Typical applications (literature; expand on general use-cases)

• Quantification of creatine in:
  • Cell/tissue lysates (metabolism studies, energy homeostasis research).
  • Food/nutrition matrices (e.g., meat extracts) after appropriate sample cleanup.
  • Bioprocess or fermentation media to monitor metabolite levels.
• Assay principles commonly used:
  • Enzymatic conversion of creatine to sarcosine and urea via creatinase/creatine amidinohydrolase, followed by sarcosine oxidase to generate H2O2; horseradish peroxidase then couples H2O2 to a chromogenic dye, yielding a colored product measurable at ~540–600 nm (literature concept; specific chemistry for this item is not specified).
  • Alternative coupled reactions or direct color formation may also be used (literature).

Practical tips (general)

• Prepare a fresh standard curve each run using known creatine concentrations spanning your expected sample range.
• Deproteinize complex biological samples where high protein may interfere or consume reagents.
• Include a sample blank (matrix matched without the color reagent or without enzyme) to correct background.
• Verify linearity by back-calculating standards; acceptable R2 is commonly ≥0.99 (literature guidance).

Reaction Conditions

Item-specific parameters (from Product Data)

• Not specified for this item; refer to CoA/Spec Sheet and kit insert for exact incubation times, temperatures, and readout wavelengths.

Typical microplate colorimetric assay conditions (literature; not item-specific)

• Temperature: Room temperature to 37°C; many enzyme-coupled assays specify 37°C to optimize kinetics.
• Incubation: 10–40 minutes until color stabilizes; kinetic mode may also be used to verify linearity.
• Detection: Visible absorbance, commonly in the 540–600 nm range for peroxidase dye systems (e.g., quinoneimine dyes); verify the exact λmax in the kit insert.
• Reaction volume: 50–200 µL per well in 96-well plates; scale appropriately for 384-well.
• Calibration: Prepare ≥5 standards across the expected range; include a reagent blank and a matrix-matched sample blank when possible.
• Interferences: Reducing agents (high DTT, βME), oxidants (H2O2 in samples), strong detergents, and azide can impair signal depending on chemistry. Protein and colored matrices may require deproteinization or dilution.

Performance checks (general)

• Acceptance criteria: Back-calculated standards within ±10% of nominal and R2 ≥0.99 are common research benchmarks.
• Plate reader setup: Ensure pathlength correction is off unless using compatible algorithms; keep consistent read height and shaking parameters.

Safety and Handling

Item-specific facts (from Product Data)

• Storage: Store at 2–8°C, protected from light.
• Shipping: Wet ice.
• GHS/Signal word/H-statements/Pictograms: Not specified for this item; refer to the SDS for each component.
• Research use: For research use only.

General safety guidance for enzyme/colorimetric assay kits (literature; consult SDS for authoritative details)

• PPE: Laboratory coat, nitrile gloves, and safety glasses are recommended. Handle all biological samples as potentially biohazardous.
• Light sensitivity: Chromogenic/fluorogenic substrates and color reagents are typically light sensitive—minimize ambient light exposure and keep bottles closed.
• Incompatibilities: Peroxidase-based systems are inhibited by azide; avoid sodium azide in samples or dilution buffers. Strong oxidants/reductants may interfere with signal development.
• Avoid contamination: Use clean, dedicated tips/reservoirs to prevent cross-well carryover and false positives.
• First aid (overview): Skin/eye contact—rinse with water for ≥15 minutes; inhalation—move to fresh air; ingestion—rinse mouth; seek medical attention as needed. Always defer to the SDS.
• Waste disposal: Collect dye-containing and peroxide-generating assay waste according to local regulations; many reagents are aqueous but may require disposal as hazardous chemical waste due to dyes or preservatives.

Good laboratory practice

• Equilibrate refrigerated components to room temperature before use unless instructed otherwise; keep enzymes on ice during setup.
• Calibrate pipettes; run blanks and standards on each plate to identify contamination or drift.

Solvent Selection

Not typically applicable. This product is a multi-component assay kit, not a single solvent or solute for reaction media.

General guidance (literature; assay context)

• Use the diluent/buffer supplied with the kit (if provided). If substitution is necessary, select a neutral pH buffer that is compatible with the assay chemistry.
• For peroxidase-coupled colorimetric assays, avoid sodium azide and strong reducing agents (e.g., DTT at high concentrations) that quench signal.
• If samples require deproteinization, common approaches include perchloric acid or TCA precipitation followed by neutralization, or ultrafiltration; ensure residual acids/salts do not exceed the tolerances of the assay (follow the kit insert).

Storage and Reconstitution

Item-specific facts (from Product Data)

• Storage conditions: Store at 2–8°C, protected from light.
• Shipping: Wet ice.

Kit-component specifics (volumes, forms, reconstitution): Not specified for this item; refer to CoA/Spec Sheet and the kit insert supplied with the product.

General guidance for assay kit storage and preparation (literature; not item-specific)

• Light-sensitive reagents (chromogens, peroxidase substrates) should remain in amber vials and be returned to 2–8°C immediately after use.
• Enzyme solutions are commonly kept cold during setup; avoid repeated freeze–thaw (if any components are frozen/lyophilized, aliquot after reconstitution as directed in the kit insert).
• Allow refrigerated components to equilibrate briefly to room temperature before opening to minimize condensation, unless otherwise instructed.
• Label aliquots with lot number, date, and component ID; do not mix components from different lots.
• Stability/expiry: Use within the labeled shelf life; once reconstituted (if applicable), observe the recommended on-board stability. For this item, these details are Not specified; refer to CoA/Spec Sheet.

Research use note

• For research use only; not for diagnostic procedures.

Structure and Identity

Item-specific facts (from Product Data)

• Product type: Creatine Content Assay Kit (Micro Method)
• SKU: C1522092
• Grade: BioReagent
• Category: Life Science (研究/实验用)
• Chemical identifiers (CAS, CID, InChIKey, SMILES): Not applicable to a multi-component assay kit; not specified for this item.

General description (literature)

• The kit is intended to quantify creatine (N-(aminoiminomethyl)-N-methylglycine) in biological or biochemical samples. As a kit, it typically contains enzymes, chromogenic/fluorogenic reagents, buffers, and a creatine standard; exact components are not specified for this item; refer to Kit Insert/CoA.
• Creatine (analyte) structural features (literature): small zwitterionic guanidino acid derivative bearing a guanidinium functionality and a carboxylate; formula C4H9N3O2; MW 131.13 g/mol (literature, for the analyte only). It is frequently present as creatine monohydrate in solid standards.
• Typical assay chemistries (literature): enzymatic conversion of creatine to intermediates culminating in H2O2 formation, which reacts with a peroxidase/dye couple to yield a measurable color or fluorescence; or direct color formation via coupled reactions. The 2D depiction would show a central carbon with methyl and carboxylate substituents and a terminal guanidino group.

Note: Structural identifiers are not applicable to the kit as a whole and are provided above only for the analyte (creatine) as literature information.

Synthetic Utility

Not applicable. This product is an assay kit designed for analytical quantification of creatine and is not used as a building block, reagent, or catalyst in synthetic chemistry.

For users seeking analytical integration in synthesis workflows (general)

• The assay can support metabolite monitoring in chemoenzymatic or cell-free systems where creatine is a process analyte; however, it does not participate in the synthesis itself.
• If you require chemical derivatization methods for creatine (e.g., for HPLC/LC–MS), consider orthogonal analytical approaches; this colorimetric kit serves as a rapid screening tool (research use only).

Target Specificity

Not applicable. This product is not an antibody or affinity reagent and does not have epitope/clone/isotype attributes.

Assay specificity (general, literature)

• Enzymatic creatine assays aim to be selective for creatine over structurally related compounds (phosphocreatine, creatinine, sarcosine). Specificity depends on the enzyme set used (e.g., creatinase vs. creatininase pathways) and is defined in the kit’s validation data.
• For this item, kit-specific cross-reactivity data are Not specified; refer to CoA/Spec Sheet.

よくある質問

What does BioReagent mean?

BioReagent indicates that the product has been processed and tested for biological and biochemical applications. Specifications may include endotoxin limits, microbial controls, sterility, low DNase/RNase/protease activity, and verified biological activity where relevant.

How should this product be stored?

Store at 2–8 °C, protected from light. The material is light-sensitive — keep it in its original opaque or amber container.

How is this product shipped?

This product ships chilled on wet ice. Unpack on arrival and transfer it to the storage condition stated above.

What documentation is provided?

Available product documentation, including Certificates of Analysis (COA), Safety Data Sheets (SDS), and specification sheets, is shown in the product document area. Document availability and access follow the current site policy.