tech1
$0.00

Fluorescence Lifetime (FLT)

FLT Probes and Labels:

Excitation Light Sources Characteristics
Product Number
(Specs Sheet)
Product Name
(Product Info)
Target Group 380 405 436 488 532 594 635 650 680 700 750 780 Medium λ abs
[nm]
ε
[M–1
cm–1]
λ em
[nm]
QY
[%]
FLT
[ns]
Buy
                   
K1-204
K1-204
SeTau-380-NHS
SeTau-380-NHS
NH2 BSA, D/P=1 383 23800 468 44 30.6
Product Number
K1-204
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K6-207
K6-207
3-DAB
3-DAB
EtOH 460 8600 665 22 4.0
Product Number
K6-207
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K7-547
K7-547
SeTau-405-NHS
SeTau-405-NHS
NH2 IgG, D/P=8 405 13800 518 19 8.4
Product Number
K7-547
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K7-567
K7-567
SeTau-405-Azide
SeTau-405-Azide
triple-CC EtOH 391 15000 498 55 8.5
Product Number
K7-567
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K8-1342
K8-1342
Seta-670-NHS
Seta-670-NHS
NH2 BSA, D/P=1.5 681 180000 696 27 2.43
Product Number
K8-1342
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K7-545
K7-545
SeTau-425-NHS
SeTau-425-NHS
NH2 BSA, D/P=1 427 4200 528 16 26.2
Product Number
K7-545
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K8-1346
K8-1346
Seta-670-Azide
Seta-670-Azide
triple-CC PB 7.4 667 180000 686 7 0.42
Product Number
K8-1346
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K8-1352
K8-1352
Square-660-NHS
Square-660-NHS
NH2 BSA D/P=1.2 676 182000 695 13 3.32
Product Number
K8-1352
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K8-1407
K8-1407
Square-650-pH-NHS
Square-650-pH-NHS
NH2 IgG, D/P=0.8
pH=2.0
662 135000 677 7 1.52
Product Number
K8-1407
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K8-1626
K8-1626
Seta-633-di-NHS
Seta-633-di-NHS
NH2 IgG, D/P=1 639 192000 650 33 3
Product Number
K8-1626
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K8-1663
K8-1663
Seta-633-NHS
Seta-633-NHS
NH2 IgG D/P=1 637 250000 647 26 1.49
Product Number
K8-1663
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K9-4119
K9-4119 new
SeTau-665-NHS
SeTau-665-NHS
NH2 IgG, D/P=1.5 662 160000 716 46 3.1
Product Number
K9-4119
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K9-4142
K9-4142
SeTau-647-di-NHS
SeTau-647-di-NHS
NH2 IgG, D/P=0.5 649 200000 691 58 3.2
Product Number
K9-4142
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K9-4148
K9-4148
SeTau-647-Maleimide
SeTau-647-Maleimide
SH PB 7.4 648 200000 692 63 3.2
Product Number
K9-4148
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K9-4149
K9-4149
SeTau-647-NHS
SeTau-647-NHS
NH2 IgG, D/P=1 648 200000 694 59 3.3
Product Number
K9-4149
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K9-4150
K9-4150
SeTau-647
SeTau-647
PB 7.4 647 211000 693 59 3.1
Product Number
K9-4150
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K7-548
K7-548
SeTau-405-Maleimide
SeTau-405-Maleimide
SH EtOH 391 15000 498 32 8.5
Product Number
K7-548
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K8-1405
K8-1405
Square-650-pH-Carboxy
Square-650-pH-Carboxy
NH2 pH 9.0 535 48000 663 9 0.53
Product Number
K8-1405
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel
K7-544
K7-544
SeTau-425-Maleimide
SeTau-425-Maleimide
SH PB 7.4 425 4200 545 39 26.2
Product Number
K7-544
Pack Size
Price
$0.00
Quantity
Total Price
$
Cancel

1. Environment-sensitive lifetime dyes

Selected Square and Seta dyes exhibit shorter fluorescence lifetimes (FLTs) in aqueous media or when labeled to low molecular weight analytes (drugs or small ligands) but increased brightness and up to ten-times longer FLTs when bound to proteins or oligonucleotides. These environment sensitive Square and Seta labels are well-suited as tracers in FLT-based assays, where e.g. the FLT-tracer would be labeled to a small antigen (ligand) displaying a short lifetime but showing a substantial FLT-increase upon binding to a larger protein (antibody or receptor). Square-660-Carboxy is also used as a fluorescent probe that binds to hydrophobic regions in cells. For a general technical note about fluorescence lifetime including additional references see FLT-TN and our product flyer on Fluorescence Lifetime Probes and Lables.

The table below provides data of the expected changes in lifetime for selected FLT dyes. It is important to note that the observed lifetime change of a fluorescently labeled ligand will strongly depend on the nature and size of the biomolecule it binds to:

 

ProductDyeDye–BSA ConjugateΔτ [ns]
τ(Dye–conj.) – τ(Dye)
τmean [ns]D/P ratioτmean [ns]
0.42 1.5 2.43 2.01
0.27 1.2 3.32 3.05
0.25 0.9 2.81 2.56

 

All of these environment-sensitive FLT labels are excitable with a 633, 635 or 647 nm laser source and Square-660 and Seta-670 can also be excited with a 405 nm laser.

 

a) FLT probes

Probes such as Square-660-Carboxy show large changes in FLT upon binding to either proteins or cell-surfaces. Square-660-Carboxy is water-soluble but with a strong affinity for the hydrophobic regions of biomolecules.

Square-650-pH is a NIR probe that exhibits a pKa around 7 and is therefore useful for measurement of pH changes in the physiologically relevant pH range from pH 6 to pH 8. Importantly both forms (protonated as well as unprotonated) of Square-650-pH are fluorescent and they exhibit different lifetimes which makes this compound useful for lifetime based applications (see Figure below). For more information about these dyes we refer you to their specs sheets and the published literature.

 

pH-ProbepHτ [ns]
Square-650-pH 2.0 1.17
9.0 0.53

 

 life1  life1
Frequency responses of Square-650-pH
at acidic and basic pH
Phase angle changes vs. pH for Square-650-pH
measured at 100 MHz

 

b) Hydrophilic FLT labels

These are reactive labels that show changes in FLT upon binding to either proteins, oligonucleotides or cell-surfaces. In general they are labeled to a low-molecular-weight binding partner and interaction with a high-molecular-weight protein or receptor leads to a change in FLT:

 life1  life1
Comparison of the frequency responses of Seta-633
before and after binding to protein or oligos

(τfree = 250 ps; τIgG bound = 1.49 ns)
Comparison of the intensity decays of Seta-633
before and after binding to protein or oligos

(τfree = 250 ps; τIgG bound = 1.49 ns)
   
K8-1351-free-and-bound-FD K8-1351-free-and-bound-TD
Comparison of the frequency responses of Square-660-NHS
before and after binding to protein or oligos

(τfree = 290 ps; τBSA bound = 3.32 ns)
Comparison of the intensity decays of Square-660-NHS
before and after binding to protein or oligos

(τfree = 290 ps; τBSA bound = 3.32 ns)

 

Applications:

Bioconjugate Chem. 20, 1807–1812 (2009).

Examples of lifetime-based assays:
- Immunoassays
- Amino-peptidase activity measurements
- Caspase-activity measurements

 

 

2. Environment-insensitive lifetime labels

a) Long-lifetime, short-wavelength FLT labels

These proprietary long-lifetime labels with lifetimes up to 32 ns in water are exitable in the UV or blue region. They are useful as polarization probes for the measurement of high MW analytes (small - medium size proteins). The measurement of BSA (MM of 65 K) in a FP assay with SeTau-425-NHS demonstrates that a suitable dynamic range can be achieved even for large antigens with  this long-lifetime polarization probe. Importantly the FLT of these labels does not change upon labeling or binding to proteins or other analytes. Several labels with fluorescent lifetimes of 32, 26 and 9 ns in aqueous environments are available.

 life1  life1

Relevant ProductsLifetime [ns]
SeTau-380-NHS 32.5
SeTau-405-NHS and SeTau-405-Maleimide 9.1-9.3
SeTau-425-NHS 26.2

 

b) Long-wavelength, squaraine-rotaxane-based FLT labels

SeTau-647-NHSSeTau-647-Maleimide and SeTau-647-di-NHS are FLT labels with lifetimes that are insensitive to the environment. The average lifetime of these labels is in the order of 3 ns which make them superior lifetime or polarization probes over Cy5 or Alexa dyes (Cy5, Alexa 647: lifetime ~ 1 ns in water). The longer lifetimes make them also more suitable donors in energy-transfer-based assays assuring that even at higher energy transfer rates the lifetime of these labels is not too short to be accurately measured with inexpensive instrumentation.

SeTau-647-maleimide has been successfully used in the studies of the mechanism of muscle contractions using FCS, specifically fluorescence lifetime correlation spectroscopy (FLCS), fluorescence lifetime and time-resolved fluorescence polarizations methods [27].

 

 life1  life1
Frequency responses of SeTau-647-NHS in
aqueous buffer solution pH 7.4
Intensity decays of SeTau-647-NHS in aqueous
buffer solution pH 7.4 in comparison to Alexa 647

 

 

Please provide following information to download software:

Your Name*:
Email ID*:
Contact Number:
Cancel