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caspase 9 activity inside whole living cells without using
antibodies.
See See Primary neuron data
prices
& sizes
read the full green procedure in the manual
read the full red procedure in the manual
see data in our newsletter
watch our apoptosis
slide show
Caspases revealed in yeast!
data
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ICT’s caspase 9 FLICA™ kits measure active caspase 9
inside whole living cells. They are NOT
ELISA kits and do NOT use antibodies.
ICT has 2 kits to detect caspase 9
- both are FLICA kits - one uses a green
fluorescent inhibitor, FAM-LEHD-FMK and another uses a
red fluorescent inhibitor, SR-LEHD-FMK.
The green FAM FLICA excites at around 490nm and
emits at 530nm;
What's
in the kits?
The FAM FLICA™kit
contains: the green caspase 9 FLICA™ probe (FAM-LEHD-FMK);
10x apoptosis wash buffer (which does not lyse the
cells); Hoechst stain (a blue DNA stain);
propidium iodide (a red live/dead stain); and a
fixative (which is used after labeling the
cells).
The SR FLICA™kit contains: the red caspase 9 FLICA™
probe (SR-LEHD-FMK); 10x apoptosis wash buffer
(which does not lyse the cells); Hoechst stain (a
blue DNA stain); and a fixative (which is used
after labeling the cells).
How
do they work?
The FLICA™ reagent (Fluorescent Labeled Inhibitors of Caspases)
is comprised of 3 segments - it includes: a green
(FAM =carboxyfluorescein) or red (SR=sulforhodamine)
fluorescent label; an amino acid peptide inhibitor sequence targeted by active
caspase 9 (LEHD); and a fluoromethylketone group (FMK) which acts as a leaving group and forms a covalent bond with the active enzyme.
The caspase 9 FLICA™ reagent
is cell permeant, so you don’t have to lyse or permeabilize the membrane to get it into the cell. Just culture your cells, add the FLICA™ probe to the media, incubate, and wash the cells. If there are active
caspase 9 enzymes, they will bind to FLICA™ and retain it within the cell, thereby capturing the fluorescent signal.
The FLICA™
probe constantly fluoresces, therefore any unbound reagent must be washed back out of the cell to remove any background noise. |
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This is done by several quick rinse and spin steps, or further incubation with the wash buffer or
cell culture media. Click here to read our
complete manual
.
How
do I read my cells?
The resulting positive fluorescent signal can be
detected with a fluorescence microscope (see newsletter
Figures 2, 3, 5a, 9), a fluorescence plate reader (Figure
10), or a flow cytometer
(Figure 7). The green FLICA™ probe, FAM, excites at 490nm and emits at
520nm; the red SR FLICA™ probe excites at 560 and
emits at 590.
How
many cells do I need?
You can detect caspase 9 activity in individual cells (Figures 1, 2, & 6) or culture them up to
1x106 cells/mL and read them using a fluorescence plate reader or flow cytometer (Figures 3 & 5). They also work with adherent cells (Figure 6).
What
kind of cells can I use?
FLICA™ kits have been used successfully with
many types of cancer cells; epithelial cells;
endothelial cells; macrophages; and other types of
human, mouse, rat tissues, and many other species.
It will not work in previously fixed
cells or paraffin cells, as that treatment
inactivates the caspase 9 enzyme. See our accessories
page for a caspase 9 antibody and other reagents.
What
other kits do you have?
To detect apoptosis in general, use the green or red poly
caspases kits. These kits use a general peptide sequence
(VAD) which reacts with all active caspases. To target
other specific caspase, ICT offers
several green FAM-FLICA™ kits which react with different caspases (caspase
1, 2, 3&7,
6, 8, 10, or 13) and additional red SR-FLICA™ kits which react with all caspases,
and 3&7. Use the red and green kits together and with our serine protease and cholinesterase kits for dual-labeling studies (Figures
5, 7, & 9).
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FLICA™
Caspase 9 Detection Kits
(green & red fluorescent inhibitors) |
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25
tests/kit |
100
tests/kit |
| Cat
# |
Price |
Cat
# |
Price |
| 9 |
Green
FAM-LEHD-FMK Caspase 9 FLICA™ Kit |
912 |
$164 |
913 |
$459 |
| 9 |
Red
SR-LEHD-FMK Caspase 9 FLICA™ Kit |
960 |
$179 |
961 |
$489 |
Order today!
Just call
1-800-829-3194
or download our simple order
sheet (Word) or PDF
main: 952-888-8788
fax: 952-888-8988
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To get your free t-shirt, just order any 2 products from
this website or our newsletter
and mention code VOL6 when placing the order.
It's that easy! (Sorry, offer does not apply to orders
placed with most distributors.)
If you have any questions about your
order, please contact Sally Hed at 800-829-3194 or
952-888-8788 or sally@immunochemistry.com
.
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References
1. Grabarek, J., P. Amstad, and Z. Darzynkiewicz. 2002. Use of fluorescently labeled caspase inhibitors as affinity labels to detect activated caspases. Human Cell vol 15(1):1-12.
2. Grabarek, J., and Z. Darzynkiewicz. 2002. In situ activation of caspases and serine proteases during apoptosis detected by affinity labeling their enzyme active centers with
fluorochrome-tagged inhibitors. Exp. Hematology vol 30:982-989.
3. Grunewald, S., Paasch, U., Said, T.M., Sharma, R.K., Glander, H.J., Agarwal, A. 2005. Caspase activity in human spermatozoa in response to physiological and pathological stimuli. Fertility and Sterility vol 83:1106-1112.
4. Paasch, U., Sharma, R.K., Gupta, A.K., Grunewald, S., Mascha, E.J., Thomas,
A.J. Jr., Glander, H.J., and Agarwal, A. 2004. Cryoperservation and thawing is associated with varying extent of activation of apoptotic machinery in subsets of ejaculated human spermatozoa. Biology of Reproduction vol 71:1828-1837.
5. inner-Staines, B., VandenHoek, S. 2004. Using cryopreserved neuronal cells for multiwell neurotoxicity assays. Poster presented at the Society for Neuroscience October 2004 annual meeting.
6. Trummell, H.Q., Raisch, K.P., Seay, L.L., and Bonner, James, A. 2004. AKT activity following exposure to IMC-C225 alone, radiation alone or a combination of both treatments. Poster presented at the American Association for Cancer Research, March 27-31, 2004, abstract # 1293.
7. VanOsten, R.L., Moore, J.M., Karacay, B. Griffith, T.S. 2005. Histone deacetylase inhibitors modulate renal cell carcinoma sensitivity to TRAIL/Apo-2L-induced apoptosis by enhancing TRAIL-R2 expression. Cancer Biology & Therapy vol 4, issue 10:31-39.1.
8. Wang, Z., Goulet, R. 3rd, Stanton, K.J., Sadaria, M., and Nakshatri, H. 2005. Differential effect of anti-apoptotic genes Bcl-xL and c-FLIP on sensitivity of MCF-7 breast cancer cells to paclitaxel and
docetaxel. Anticancer Res vol 25(3c):2367-2379.
See all FLICA
references (pdf)
Order today!
Just call
1-800-829-3194
or download our simple order
sheet (Word) or PDF
main: 952-888-8788
fax: 952-888-8988
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