Product manual
Product use, preparation, operating conditions and storage.
FAMReactive dye
5-FAM free acid
Cat. No.QH-60-6006
A free-carboxylic-acid form for custom coupling after carboxyl activation or fluorescence-method development.
Packs & quoteCOOH
Carboxyl-functional dyeCO–X
Activated carboxylCO–NH–R
Amide linkageExcitation488 nm
Emission525 nm
TargetPrimary amines coupled through a suitable activation route
Carboxyl activation and coupling
A free carboxylic acid is not a preactivated NHS ester. Activate it using a sample-compatible method before coupling to an accessible primary amine; do not use it as an NHS reagent without activation.
Custom fluorescent conjugation
For small molecules, peptides or amino-modified samples with a defined activation, coupling and purification procedure.
Analytical method development
When comparing free dye and conjugate detection, match pH, solvent and concentration and consider environment-dependent spectral differences.
Specifications
Product information
- Product name
- 5-FAM free acid
- Cat. No.
- QH-60-6006
- Product form
- Free acid
- Target
- Primary amines coupled through a suitable activation route
- Pack sizes
- 1 g
Preparation and properties
- Excitation wavelength
- 488 nm
- Emission wavelength
- 525 nm
- CAS
- 76823-03-5
- Molar extinction coefficient
- 83,000 M⁻¹ cm⁻¹
- Quantum yield
- 0.93
- Storage
- Store at -20°C, protect from light and moisture
Spectral information and detection
Product parameters
Use parameters for the exact catalog number when preparing reagents, setting reagent input or calculating results. Request a lot-specific CoA for test results.
Protocol
Activation scheme
Choose activator, solvent and pH for the actual carboxylic acid and target amine, then begin at small scale.
Limit off-target reactions
For targets containing multiple carboxyls and amines, assess self-crosslinking, site distribution and separation difficulty.
Isomer and reactive form
Record the 5- or 6-isomer and free-acid or NHS-ester form. Wavelengths differ among isomers and forms; retain the exact catalog number during lot comparisons, HPLC separation and probe transfer.
Activate the dye carboxyl
Choose an activation system compatible with this dye carboxylic acid and solvent. Record activator input, pH and time; do not add unactivated free acid to the sample as though it were an NHS ester.
Couple to the target primary amine
React the activated dye with a sample carrying accessible primary amines. For samples with multiple carboxyl and amino groups, compare separate activation and excess-activator removal, monitoring self-crosslinking and recovery.
Purification and analysis
Select desalting, size-exclusion separation, dialysis or chromatography for the sample size. Remove free reagent before measuring concentration, labeling density and function. Residual free dye raises background and biases labeling-density calculations.
Confirm product composition
Use suitable chromatography, mass spectrometry or electrophoresis to distinguish conjugate, unreacted dye and by-products, then test probe function. Increased fluorescence or a color change alone does not establish covalent coupling.
Request product documentation
Select a document and enter your phone and email. Include the lot number for a CoA.
5-FAM free acidQH-60-6006
Spectra and measurement conditions
Spectra, measurement conditions and detection settings for the exact catalog number.
Certificate of analysis (CoA)
Lot-specific test results. Include the lot number from the vial.
Safety data sheet (SDS)
Handling, storage and emergency response information.
Quality and lot information
Unactivated-dye control
Omit the carboxyl activator, mix an equal amount of free dye with the sample and purify identically to assess adsorption and free-dye carryover.
Unlabeled sample
Use an unlabeled aliquot from the same preparation to assess changes in function, aggregation and recovery.
Free dye and negative sample
Include free-dye or no-target-handle controls to distinguish conjugation from adsorption. Match exposure, gain or voltage during detection.
Free dye and probe quality
Use suitable HPLC, PAGE or mass spectrometry to assess probe purity and labeling, comparing function with the unlabeled sequence. Measure free dye and probe in the same buffer and pH so environmental changes are not mistaken for changes in labeling.
Technical articles
Application support
Include your sample, formulation and detection method when requesting application guidance.
References
EDC/NHS carboxyl-to-amine coupling
Thermo Fisher Scientific · Carbodiimide crosslinker chemistry
Explains carboxyl activation, intermediate hydrolysis and self-crosslinking risks in multi-handle samples, informing free-acid dye activation and product analysis.
Frequently asked questions
Can it replace the corresponding NHS ester?
Not directly. The free acid needs activation; use this product’s molecular weight, solubility and input conditions.
Can it directly stain nuclei?
A fluorescent free acid is used for analytical or coupling work; its fluorophore does not make it a specific nucleic-acid stain.
How is conjugation confirmed?
Remove free dye, characterize the product by suitable chromatography, mass spectrometry or electrophoresis, and check sample function.