AnaSpec’s
SensoLyte® MMP assay kits, especially the SensoLyte® 520
MMP Assay Kits, have become the new standard in MMP assay sensitivity.
With published results in world-class journals and the industry’s longest wavelength,
the SensoLyte® 520 line is one of the most sensitive
(Figure 1) and brightest lines of MMP assay kits in the world.
SensoLyte®
520 MMP Assays
• Measure MMP activity
• 96-well or 384-well format
• Mix and read
• Fluorimetric - long wavelength
• 5-FAM/QXL™ 520 FRET pair (Ex/Em=490/520 nm)
• Minimal autofluorescence
• Assay time – as short as 1h
 |
| Figure 1. The performance of an identical peptide sequence,
one coupled with 5-FAM/QXLTM 520 and the other coupled with EDANS/DABCYL
FRET pairs was compared (A). Fluorescence was measured after
1 hr incubation of FRET substrates with serial dilutions of MMP-13 (Flex800,
Bio-Tek instruments). A 5-FAM/QXLTM 520 MMP substrate was compared
with Mca/Dpa FRET substrate (B). Fluorescence was measured after 1 hr incubation
of substrates using serial dilutions of MMP-13 (Flexstation 384II, Molecular
Devices). |
SensoLyte®
Plus 520 MMP Assays
• Measure MMP activity
• Enhanced specificity – human capture antibody
included
• 96-well format (strips)
• Mix and read
• Fluorimetric - long wavelength
• 5-FAM/QXL™ 520 FRET pair (Ex/Em=490/520
nm)
• Minimal autofluorescence
SensoLyte®
490 MMP Assays
• Measure MMP activity
• 96-well or 384-well format
• Mix and read
• Fluorimetric
• EDANS/DABCYL FRET pair (Ex/Em=340/490 nm), except
71132 NBD/DMC FRET pair (Ex/Em=370/460 nm)
• Assay time – as short as 1h
SensoLyte®
MMP ELISA Assays
• Measure MMP amount
• 96-well format
• Colorimetric (Absorbance)
Also available:
SensoLyte® 520 Generic MMP Assay Kit *Fluorimetric*
SensoLyte®
520 MMP Substrate Sampler Assay Kit *Fluorimetric*
SensoLyte®
520 MMP Profiling Assay Kit *Fluorimetric*
MMP Recombinant
Proteins
• 100 ul (10 ug/ml)
• Highly active after APMA incubation
• Serve as positive controls
for SensoLyte® MMP assay kits
• Can also be used for zymography, Western blot, IP, and ELISA
Anti-MMP
Antibodies
• Human, zebrafish
(Z-Fish™), mouse and rat specific
• Epitopes from hinge,C-terminal (CT), N-terminal
(NT), IN (internal) regions
Posters
Presented at Scientific Meetings:
Table. Publications
citing the use of AnaSpec’s SensoLyte® MMP assay kits and their applications.
| Cat# |
Product |
Application |
| 71150 |
SensoLyte™
520 MMP-1 Assay Kit |
MMP-1 activity measured in hyperhomocysteinemic
rats.1 |
| 71151 |
SensoLyte™
520 MMP-2 Assay Kit |
(human) pro-MMP-2 … first activated with 1 mm APMA
for 20 min or 2 h…2, 3
…tissue extracts were obtained from nonpregnant, pregnant, and
postpartum mice…and a fluorescently labeled substrate specific for MMP2
was used to determine enzyme activity.4
WT, Fbln3–/–, and Fbln5–/–
mice were anesthetized and perfused with 10% neutral buffered
formalin (pH 7.4). Thereafter, the female urogenital tract
was dissected en bloc…Vaginal tissues were thawed on
ice, minced, washed in PBS, and homogenized in MMP2 assay buffer
(EnzoLyte 520 MMP-2 assay kit; AnaSpec, San
Jose, CA)…5
Rat aortic SMCs(smooth muscle cells) and adventitial FBs (fibroblasts)
and MFBs (myofibroblasts) were obtained…The MMP-1 and MMP-2
activities in the conditioned media of control and 1-h flow-
and 6-h flow-treated gels were determined using the SensoLyte
Plus 520 MMP-1 Assay Kit and SensoLyte 520 MMP-2 Assay Kit
from AnaSpec (San Jose, CA).6 |
| 71155 |
SensoLyte™
520 MMP-9 Assay Kit |
(human) … pro-MMP-9 … first activated with
1 mm APMA for 20 min or 2 h…2, 3 |
| 71156 |
SensoLyte™
520 MMP-13 Assay Kit |
MMP-13 (rat interstitial collagenase) activity was
detected at 0 and 7 days using the SensoLyte™ (formerly Enzolyte) 520
MMP-13 assay kit.7 |
| 72017 |
SensoLyte
Plus™ 520 MMP-9 Assay Kit |
MMP-9 activities of cell [MiaPaCa2 and PANC1 human
pancreatic ductal adenocarcinoma cells]or tumor lysates were
assessed using the SensoLyte™ (formerly Enzolyte) Plus 520 Enhanced Selectivity
MMP-9 activity.8
… genetically altered cells (esophageal squamous) were
used to detect MMP-9 enzymatic activity in duplicate using the SenzoLyte
Plus MMP-9 assay kit from AnaSpec.9
Rat primary microglia (5 × 106 cells/mL) were
treated with LPS/PMA in the presence or absence of MMP inhibitors for
24 h and the supernatants were collected for measuring MMP activities.
Both the total and active MMP-3 or MMP-9 activities were assayed according
to SensoLyteTM 520 MMP-3 or SensoLyte PlusTM 520
MMP-9 assay system, respectively (AnaSpec, San jose, CA, USA). 10 |
| 72019 |
SensoLyte
Plus™ 520 MMP-13 Assay Kit |
MMP-13 activity (in RKO- human colorectal
carcinoma cell line) cells using the SensoLyte Plus™ 520 MMP-13 Assay.11
Adult human mesenchymal stem cells were encapsulated in pure PEG
hydrogels, without peptide functionality, at a concentration of 2 × 106 cells/ml…
Cell/polymer constructs were removed from culture at specific time points
and analyzed for production of MMP-13 via the SensoLyte Plus™ (formerly
EnzoLyte Plus™) 520 MMP-13 Assay Kit (AnaSpec).12
Serum-free D-MEM (Dulbecco's modified Eagle medium) conditioned
by confluent stromal cell cultures for 24 h was concentrated approximately
40 fold…and analyzed using SensoLyte Plus 520 MMP-1 and SensoLyte Plus
520 MMP-13 Assay Kits (AnaSpec; San Jose, California, USA).13 |
| 71158 |
SensoLyte™
520 Generic MMP Assay Kit |
General MMP activity (from rat heart) was measured
using the SensoLyte™
(formerly Enzolyte) 520 Generic MMP assay kit.14
General MMP activity was assessed using the EnzoLyte™ 520 Generic
MMP assay Kit (AnaSpec, San Jose, CA). This kit is optimized to detect
the activity of different MMPs such as MMP-1, -2, -3, -7, -8, -9, -12,
-13 and -14, using a fluorescence-quenched substrate (5-FAM/QXL™520).15
The SensoLyte (formerly EnzoLyte) 520 Generic MMP Fluorometric
Assay Kit (AnaSpec) was used according to the manufacturer's
directions. Cell culture supernatants were incubated with 1
mmol/L of 4-aminophenylmercuric acetate for 24 h at 37°C to
activate pro-MMPs. 16
WT mice of the genetic background C57BL/6 were obtained from
Jackson Laboratories…MMP activity was determined using the SensoLyte
520 Generic MMP Assay Kit (AnaSpec) as per manufacturer's
instructions. Briefly, media from WT, TIMP-3–/–,
and WT cells treated with rTIMP-3 (1 µg/ml) were collected
72 h postseeding, and 50 µl of these media were incubated with
the FAM/QXL 520 fluorescence resonance energy transfer substrate
for 1 h in a black 96-well plate at room temperature in the
dark. 17
Thirty human fibroblast populations were isolated from the skin
biopsies of SSc(Systemic sclerosis) patients…Total MMP activity was determined
in the same supernatant as that for collagen assay using the SensoLyteTM
520 Generic MMP assay kit (Anaspec, San Jose, CA, USA). This kit can detect
simultaneously the activity of MMP-1, 2, 7, 8, 9, 12, 13 and 14.18 |
| 71170 |
SensoLyte™
520 MMP Substrate Sampler Kit |
Aspergillus culture supernatants, human serum and the
mixture of both..(using)..fluorescence resonance energy transfer (FRET)
were chosen out of a commercially available protease assay kit comprising
16 different substrates SensoLyte™ (formerly Enzolyte) 520 matrixmetalloprotease
(MMP) substrate sample kit, AnaSpec.19
The activity of MMPs in the conditioned medium and cell
lysates (CHO or APP-overexpressed neuroblastoma cells) was determined
using SensoLyte™ (formerly EnzoLyte) MMP fluorometric
assay kit.20
Murine Metastatic 4T1 and normal NmuMG cells were obtained…The
activated samples were assay for MMP-9 activity using the SensoLyte 520
MMP Sampler Kit (AnaSpec).21 |
| 72025 |
SensoLyte® 520 MMP-14
Assay Kit |
To evaluate MT1-MMP activity, we prepared membrane
fractions of HAECs (human aortic endothelial cells)…cells
were lysed with a hypotonic buffer, then sonicated and centrifuged
at 15 000 g for 10 min. The separated membrane fractions
were assessed by the commercially available fluorescent assay
kit (SensoLyte 520 MMP-14 assay kit, AnaSpec,
San Jose, CA, USA) according to the manufacturer's instructions.
22 |
| 72012 |
SensoLyte®
Plus 520 MMP-1 Assay Kit |
Rat aortic SMCs(smooth muscle cells) and adventitial
FBs (fibroblasts) and MFBs (myofibroblasts) were obtained…The
MMP-1 and MMP-2 activities in the conditioned media of control and
1-h flow- and 6-h flow-treated gels were determined using the
SensoLyte Plus 520 MMP-1 Assay Kit and SensoLyte 520 MMP-2 Assay
Kit from AnaSpec (San Jose, CA). 6
Primary cell cultures were established from biopsy and resection
specimens from patients pre-operatively diagnosed with GCT…Serum-free
D-MEM conditioned by confluent stromal cell cultures for 24 h …analyzed
using SensoLyte Plus 520 MMP-1 and SensoLyte Plus 520 MMP-13 Assay Kits
(AnaSpec; San Jose, California, USA). 23 |
| 71152 |
SensoLyte®
520 MMP-3 Assay Kit |
Rat primary microglia (5 × 106
cells/mL) were treated with LPS/PMA in the presence or absence of MMP
inhibitors for 24 h and the supernatants were collected for measuring
MMP activities. Both the total and active MMP-3 or MMP-9 activities were
assayed according to SensoLyteTM 520 MMP-3 or SensoLyte PlusTM
520 MMP-9 assay system, respectively (AnaSpec, San Jose, CA, USA).10
Astrocytes were prepared from the cerebrum of 1–2-day-old Wistar
rats…Stromelysin-1 activity in the astrocytic culture medium was determined
using a stromelysin-1 (MMP-3) activity assay kit (SensoLyte 520 MMP-3
assay kit, AnaSpec, San Jose, CA).24 |
| 60580 |
520
MMP FRET Substrate XIII |
The activity of MMP-3 (stromelysin-1) (of
rat brain homogenates) was measured fluorometrically using a 5-FAM/QXL520
FRET peptide…25 |
References:
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2. Johnson,
SL. et al. Chem. Bio. Drug Design 71, 131 (2008).
3. Johnson,
SL. et al. Bioorganic Chem. 35, 306 (2007).
4. Wieslander,
C. et al. Biol. Reprod.. 10.1095/biolreprod.107.063024 (2007).
5. Rahn,
D. et al. Am. J. Pathol. 174, 206 (2009).
6. Shi,
Z-D. et al. Am. J. Physiol. Heart Circ. Physiol. 10.1152/ajpheart.00369
(2009).
7. Arnoczky,
SP. et al. Am. J. Sports Med. 35, 763 (2007).
8. Liau, S-S. et
al. Cancer Res. 66, 11613 (2006).
9. Okawa,
T. et al. Genes & Dev. 21, 2788 (2007).
10.
Woo, MS. et al. J. Neurochem. doi:10.1111/j.1471-4159.2008.05430.x (2008).
11. Bai,
L. et al. FEBS Lett. 581, 5904 (2007).
12. Salinas,
CN. and KS. Anseth Biomaterials 581, 5904 (2007).
13. Cowan,
RW. et al. Bone 44, 865 (2009).
14. Errami, M.
et al. J. Pharma. Exp. Ther. DOI: 10.1124/jpet.107.133975 (2007).
15. Gueders,
MM. et al. Biochem. Pharma. 75, 514 (2008).
16. Santisteban,
M. et al.Cancer Res. 69, 2887 (2009).
17. Hammoud,
L. et al. Am. J. Physiol. Cell Physiol. 296, C735 (2009).
18. Corriveau,
MP. et al. J. Pathol. 217, 534 (2009).
19. Schaal,
R. et al. J. Microbio. Meth. 71, 93 (2007).
20. White, AR. et al. J.
Biol. Chem. 281, 17670 (2006).
21. Allington,
TM. et al. FASEB J doi: 10.1096/fj.09-138412 (2009).
22. Sugimoto,
K. et al. Cardiovasc Res. doi:10.1093/cvr/cvp177 (2009).
23. Cowan,
RW. et al. Bone 44, 865 (2009).
24. Koyama,
Y. and T. Tanaka BBRC doi:10.1016/j.bbrc.2008.04.064 (2008).
25. Candelario-Jalil,
et al. J. Pharma. Exp. Ther. 323, 488 (2007).
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