FITC-avidin

Figure 7.6 Intensity decays for FITC-Avidin on both glass and SiFs, both before and after exposure to low power microwave heating. The intensity decays on the SiFs are almost identical. IRF Instrument Response Function. Adapted from reference 1.

AgNPs FITC avidin/biotinlyated BSA orHSA Geddes et al. (2006) ... 

First layer detection Add 100 pL to each slide. FITC-Avidin DCS at 1 200 dilution in 4X SSC-TB for 45 min at room temperature. 

Human Papilloma Virus Physiological state applications Solid biopsy material Biotin-nucleic acid probe -t FITC-avidin Butor et al. 1997... 

Although FITC and other reactive fluorescein derivatives still are widely used to label (strept)avidin and other proteins, better fluorescence yield and stability will be obtained if one of the newer hydrophilic fluorescein dyes is used. See Chapter 9, Section 1, for additional details on labeling proteins with fluorescein. 

Add 50-100 pi of the FITC solution to each ml of the (strept)avidin solution. 

Figure 23.6 The reaction of FITC with avidin produces a fluorescent probe via isothiourea bonds.

NPH HCl) with l-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC HCl) as a coupling agent [593,594], The advantage is that with hydrazines it is possible to carry out the derivatization in milder conditions. An efficient postcolumn derivatization is obtained using avidin or streptavidin. In fact, these two proteins react in a very specific way with biotin, therefore they are bound with a fluorescent marker, such as fluorescein 5-isothiocyanate (FITC) to obtain fluorescent derivatives. Some authors [595] report the use of MS or MS/MS for biotin detection, but this method seems to be less sensitive than FLD. 

X4 i=520nm) post column reagent avidin-FITC in phosphate buffer solution... 

Add 100 pL of TBST containing 1 ng/mL avidin-FITC, and incubate for 30 min at room temperature... 

Fluorescein isothiocyanate (FITC) is the most common fluorescent label used to modify proteins and other biomolecules (Chapter 8, section 1.1). The isothiocyanate group reacts with amines in protein molecules to form a stable thiourea linkage (Fig. 365). Avidin or streptavidin may be tagged with this reagent to yield highly fluorescent... 

Add 50—100 p,l of the FITC solution to each milliliter of the avidin or streptavidin solution. 

The first aptasensor reported was particularly based on optical detection [66]. The 58-mer RNA aptamer selective to L-adenosine was immobilized onto the core of multimode fiber using avidin-biotin method. The detection was based on competitive binding of FITC-labeled L-adenosine with unlabeled analyte. This sensor also allowed to study the kinetics of binding and determine equilibrium constants. 

An avidin-biotin system has been used to attach antibodies in the bilayer of DDSs. Xiao et al. developed a three-step strategy to improve the tumor-to-tissue ratio of anticancer agents [184], Two antibodies specific for the CA-125 antigen that is highly expressed on NIH OVCAR-3 cells were used. These cells were prelabeled with biotinylated anti-CA-125 antibody and fluoroscein isothiocyanate (FITC)-labeled streptavidin (SAv) prior to administration of biotinylated liposomes. Both antibodies were specifically bound to the cell surface of OVCAR-3 cells but not to SK-OV-3 cells, which do not express the specific antibody. Antibody biotinylation did not affect its immunoreactivity. 

Avidin-FITC stock (1000X) obtained from Sigma. Made as a 5 mg/mL solution in PBS. This stock solution should be aliquoted and stored at -20°C. Working stock solutions can be stored at 4°C but should be used within 1 mo and should only be diluted to a working dilution when required. 

Avidin-FITC solution 4X SSC (diluted from 20X SSC using dH20) + 0.1% (v/v) Triton X-100 and 5% (w/v) fat-free dried skimmed milk (Marvel, Premier Beverages, Stafford, UK) +5 pg/mL avidin-FITC (purchased from Sigma). Makeup fresh for each experiment. 

Fig. 1. Incorporation of bt-dUTP by exogenous TdT into the DNA of human ovarian carcinoma cells exposed to the DNA damaging agent cisplatin. Two-dimensional frequency contour plots of red fluorescence (x-axis PI stained DNA) vs green fluorescence (y-axis, bt-dUTP labeled with avidin-FITC). Untreated human A2780 ovarian carcinoma cells these cells were exposed to 20 pM cisplatin for 1 h and then incubated for 96 h at 37°C. They were then processed using Method B. Cells with green fluorescence above the dotted line are considered to be positive for incorporation of labeled nucleotide by exogenous TdT and therefore undergoing apoptosis.

Resuspend the pellet in 50 pL of the avidin-FITC solution containing 5 pg/mL avidin-FITC (1 in 1000 dilution of the stock solution). Incubate in the dark at room temperature for 30 min. 

Probe Detection Using Avidin and Antibodies for Two Colors (FITC/Texas Red)... 

Butor et al. (1997) were able to detect and map Human Papilloma Virus (HPV) infected cells labelled by means of FISH in cervical condyloma biopsies using a cDNA probe conjugated with biotin and subsequent detection with FITC conjugated avidin. 

---