FAM: Free acid, NHS Ester. Select the form for the sample handle and readout.
3 products
3 matching products
| Product / Cat. No. | Chemistry | Excitation / nm | Emission / nm | Pack sizes | Product details |
|---|---|---|---|---|---|
| 5-FAM NHS EsterQH-60-6007 | NHS Ester | 488 | 515 | 100 mg | |
| 5-FAM free acidQH-60-6006 | Free acid | 488 | 525 | 1 g | |
| 6-FAM free acidQH-60-6008 | Free acid | 489 | 523 | 1 g |
Need help with reactive forms or detection channels?
QH-60-6007
| Product / Cat. No. | Form | Ex / Em · nm |
|---|---|---|
| 6-FAM free acidQH-60-6008 | Free acid | 489 / 523 |
| 5-FAM NHS EsterQH-60-6007 | NHS Ester | 488 / 515 |
| 5-FAM free acidQH-60-6006 | Free acid | 488 / 525 |
6-FAM free acid; 5-FAM NHS Ester; 5-FAM free acid have defined positional-isomer and activation identities. Track starting materials and products by isomer and free-acid or NHS-ester form, with separate molar calculations.
Primary amines coupled through a suitable activation route
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.
Protein N-termini, lysine residues and introduced primary amines
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.
NHS esters address accessible primary amines; free acids require activation. For nucleic-acid probes, confirm an amino handle, then assess full-length product, free-dye carryover and probe function after purification.
Selection criteria
Compare each product’s excitation and emission with the instrument’s light source, filters and detector. A number in a product name does not replace its specifications.
Select the isomer and form for the intended probe or conjugate. Use molecular weight and optical values for the exact catalog number rather than one value for the entire dye name.
Compare specific signal, negative background and function at matched sample input and acquisition settings. Titrate the purified conjugate.
Sample and experimental preparation
Choose a suitable activation, coupling and separation scheme. Distinguish starting dye, activated intermediate and target product; assess self-crosslinking when the target contains multiple carboxyls and amines.
Record target molecular weight, concentration, available handles and buffer additives. Select a method that separates the target product from free dye, using the product conditions for the actual reactive form.
Retain an unlabeled aliquot from the same preparation. Measure concentration, labeling density and function after purification. Use the actual conjugate for single-color references in multicolor detection.
Distinguish free acids from NHS esters, plan carboxyl activation, amine coupling and product analysis, and retain positional-isomer identity for FAM and ROX.
Compare labeling density, sample recovery, specific signal and background after purification, rather than total fluorescence alone.
Tris and glycine consume NHS dye. BSA and gelatin require separation from the target protein; routine buffer exchange is insufficient.
Check NHS hydrolysis and amine-containing additives, then distinguish low labeling from sample loss or reduced binding.
Calculate DOL with absorbance correction, distinguish input ratio from attached dye, and assess recovery and binding.
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.
thermofisher.com
Reactive handles, buffer compatibility and method guidance.
Nanda JS, Lorsch JR. · Methods in Enzymology · 536:87–94 · 2014
Uses FAM ester labeling of a protein to explain primary-amine chemistry and labeling-efficiency assessment. It informs NHS workflow design; input and reaction conditions depend on the actual dye and sample.
Vira S, Mekhedov E, Humphrey G, Blank PS. · Analytical Biochemistry · 402(2):146–150 · 2010
Uses FITC-labeled anti-HA antibody to assess changes in labeling density and antigen binding. It informs input-series design and post-purification functional testing; DOL alone is insufficient for selecting a conjugate.
Request the manual, SDS or lot-specific CoA by catalog number. Include the vial lot number for a CoA.
Product use, preparation, operating conditions and storage.
Lot-specific test methods and results.
Handling, storage and emergency response information.
Identify the sample handle or required recognition mechanism, then select a matching reactive dye or preconjugated probe. Use preparation, reaction and storage requirements for the exact catalog number.
A free acid is not preactivated and needs a separate activation step. Confirm molecular weight, solubility and product analysis for each form.
Match target concentration, acquisition settings and sample processing. Compare labeling density together with specific signal, negative background, recovery and function; a free-dye or single endpoint measurement does not assess every performance characteristic.
Use molecular weight, an appropriate extinction coefficient and protein-absorbance correction for the exact catalog number after removing free dye. Request parameters not shown on the product page through its Documents tab.
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