Product manual
Product use, preparation, operating conditions and storage.
FAMReactive dye
5-FAM NHS Ester
Cat. No.QH-60-6007
Label proteins, peptides and amino-modified molecules by coupling the NHS ester to accessible primary amines.
Packs & quoteNHS
Activated esterR–NH₂
Primary amineCO–NH–R
Amide linkageExcitation488 nm
Emission515 nm
TargetPrimary amines on amino-modified nucleic acids, or accessible amines on proteins/peptides
Primary-amine coupling
The NHS ester forms an amide bond with a primary amine; hydrolysis competes with coupling. Proteins usually contain multiple accessible sites, and the input ratio is not the final degree of labeling.
Amino-modified oligonucleotides
Post-synthetically label through a defined amino handle. After purification, distinguish full-length labeled product, unreacted oligonucleotide and free dye, then assess hybridization or amplification function.
Proteins and peptides
For accessible-amine coupling, optimize input and assess function separately. A FAM or ROX fluorophore does not itself provide nucleic-acid-binding specificity.
Specifications
Product information
- Product name
- 5-FAM NHS Ester
- Cat. No.
- QH-60-6007
- Product form
- NHS Ester
- Target
- Primary amines on amino-modified nucleic acids, or accessible amines on proteins/peptides
- Pack sizes
- 100 mg
Preparation and properties
- Excitation wavelength
- 488 nm
- Emission wavelength
- 515 nm
- CAS
- 92557-80-7
- 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
Buffer
Avoid competing primary amines such as Tris, glycine and free amino acids. Remove carrier proteins such as BSA when they are not intended for labeling. Use a primary-amine-free buffer compatible with the sample.
Dye and input calculation
Prepare the activated ester using a solvent supported for this product. Convert sample and dye amounts to moles using their molecular weights and record the final organic-solvent fraction.
Verify the nucleic-acid amino handle
For post-synthetic oligonucleotide labeling, verify introduced amino handles, their number and protection state, and calculate input from the sequence molecular weight. Direct staining procedures for nucleic-acid-binding dyes do not apply to this reactive reagent.
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.
Exchange the sample buffer
Exchange into a suitable buffer and check concentration, purity, pH and visible aggregation.
Set up a reaction series
Use a small sample to compare dye input ratios. Mix gently, protect from light and record pH, temperature and time. Select conditions from this product’s instructions and sample tolerance.
Purify for the sample type
For oligonucleotides, use suitable HPLC, PAGE or another validated method to resolve full-length labeled product, unreacted nucleic acid and free dye. Select protein cleanup separately by molecular size; protein desalting conditions do not automatically retain short nucleic acids.
Verify composition and sequence function
Quantify nucleic acid with sequence-appropriate 260 nm extinction data and dye absorbance correction, not a protein A280 labeling formula. Verify labeling composition by separation or mass spectrometry and compare hybridization, amplification or other function with the unlabeled sequence. Evaluate protein conjugates using protein-specific parameters.
Request product documentation
Select a document and enter your phone and email. Include the lot number for a CoA.
5-FAM NHS EsterQH-60-6007
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
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.
Input titration
Compare a molar input series and assess signal, negative background, product recovery and function together.
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.
References
Thermo Fisher · Amine-reactive chemistry
thermofisher.com
Distinguishes amine-reactive forms such as NHS and isothiocyanate and competing buffers; separately establish whether a nucleic-acid sample has accessible amines.
Frequently asked questions
Can it directly label unmodified nucleic acids?
Routine NHS coupling requires an accessible primary amine. Confirm a suitable amino modification on the oligonucleotide.
Is a higher degree of labeling better?
Overlabeling can affect binding, solubility or signal. Assess specific signal, background and function after purification.
Can aqueous stock be stored for extended periods?
Activated esters hydrolyze in water. Use only the stock solvent, concentration and storage conditions supported for this product.
Can a protein A280 formula quantify oligonucleotide labeling?
Do not transfer it directly. Nucleic acids require sequence-appropriate A260 extinction data and dye correction. Separate free dye and unreacted nucleic acid first, then assess purity and labeling composition.