Product manual
Product use, preparation, operating conditions and storage.
Cy3Bioconjugate
Cy3-Streptavidin
Cat. No.QH-15-1014
Detect biotinylated antibodies, nucleic acids or other biotinylated molecules with fluorescent streptavidin.
Packs & quoteStreptavidin
Affinity recognitionBiotin
Biotinylated targetBinding
Noncovalent bindingExcitation554 nm
Emission564 nm
TargetAccessible sample-bound biotin
Preparation and storage
- Preparation
- Aqueous formulation
- Storage
- Store at 2–8°C, protect from light. Do not freeze
Biotin-affinity detection
Streptavidin binds biotin noncovalently. Free biotin can occupy binding sites, endogenous biotin can cause background, and multivalent binding can crosslink targets.
Biotinylated-antibody detection
Pair with a suitable biotinylated antibody for flow cytometry or immunofluorescence, checking channels and biotinylation density.
Nucleic-acid and affinity detection
Detect biotin-tagged probes or surfaces, using washing to control unbound conjugate and nonspecific adsorption.
Specifications
Product information
- Product name
- Cy3-Streptavidin
- Cat. No.
- QH-15-1014
- Product form
- Streptavidin
- Target
- Accessible sample-bound biotin
- Pack sizes
- 1 mg / 5 mg / 10 mg / 25 mg / 100 mg
Preparation and properties
- Excitation wavelength
- 554 nm
- Emission wavelength
- 564 nm
- Molar extinction coefficient
- 150,000 M⁻¹ cm⁻¹
- Quantum yield
- 0.31
- Preparation and sample
- Aqueous formulation
- Storage
- Store at 2–8°C, protect from light. Do not freeze
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
Remove free biotin
Remove excess small-molecule reagent after biotinylation. Check media and sample additives for biotin before affinity detection.
Handle the formulation
Check concentration, buffer and storage for this formulation. Handle PE, APC and tandem conjugates as fluorescent proteins, avoiding organic solvents and unsuitable freeze–thaw cycles.
Prepare the biotinylated sample
Confirm biotin incorporation, removal of free reagent and sample integrity.
Titrate the conjugate
Compare probe amounts and incubation times at matched sample input, protected from light.
Wash and detect
Remove unbound reagent and use single-stain controls for channels, compensation or unmixing before assessing signal and background.
Request product documentation
Select a document and enter your phone and email. Include the lot number for a CoA.
Cy3-StreptavidinQH-15-1014
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
Nonbiotinylated sample
Use an identically handled nonbiotinylated sample to assess nonspecific binding and endogenous-biotin background.
Single-stain and lot controls
Use this reagent for single-stain controls. Match tandem reference controls to the actual lot and processing conditions.
Probe titration
Select the amount based on negative background and positive separation, limiting nonspecific signal from excess reagent.
Technical articles
Effects of free biotin on detection
Free biotin occupies streptavidin-binding sites and can reduce detection of biotinylated targets.
For prepared streptavidin probes; check competition from free biotin.
Analysis of endogenous biotin and nonspecific background
Use unstained, streptavidin-only and non-biotinylated-probe controls to distinguish autofluorescence, endogenous biotin and adsorption.
For biotin-affinity detection; distinguish endogenous biotin from nonspecific adsorption.
Streptavidin fluorescence detection and multivalent binding
Multivalent streptavidin can crosslink biotinylated targets, affecting live-cell motion and clustering measurements.
For assessing probe-induced crosslinking; the cited engineered monovalent protein does not establish that current products are monovalent.
Application support
Include your sample, formulation and detection method when requesting application guidance.
References
Thermo Fisher · Biotin binding and interference
Thermo Fisher Scientific
Sample preparation, free biotin and background control in biotin-affinity detection.
Thermo Fisher · Fluorescent streptavidin methods
Thermo Fisher Scientific
Sample preparation, free biotin and background control in biotin-affinity detection.
A monovalent streptavidin with a single femtomolar biotin binding site
Howarth M, Chinnapen DJF, Gerrow K, et al. · Nature Methods · 3:267–273 · 2006
Uses an engineered monovalent protein to investigate crosslinking by multivalent binding. It informs assessment of probe valency in membrane-protein motion and clustering studies.
DOI: 10.1038/nmeth861
Frequently asked questions
Why is signal weak after biotinylation?
Check residual free biotin, handle accessibility, sample loss and probe detection settings.
Can it directly label a nonbiotinylated antibody?
It does not replace a reactive dye. The target needs accessible biotin for specific affinity detection.
Can endogenous biotin be ignored?
Endogenous background varies among tissues and cells. Assess it with a no-biotinylated-probe control.