ALEXIS Biochemicals: Monosodium urate (crystals)

ALX-400-047

Revised 24-Oct-07

Monosodium urate (crystals)
SYNONYMS	MSU (crystals) Uric Acid (crystals)
PRODUCT LINE	Inflammation
PRODUCT CATEGORY	Inflammasome

Ordering Information

Product Numbers:	Format:	Size:	Unit Price:	Quantity:	Add To Cart
ALX-400-047-M002		2 mg	90.00 USD

Product Specification

FORMULA:	C₅H₄N₄O₃ . Na
MW:	168.1 . 23.0
CAS NUMBER:	1198-77-2
RTECS:	YU7062000
CONCENTRATION:	5mg/ml after reconstitution.
PURITY DETAIL:	Not determined. Degradation of the crystals with uricase (which breaks purines down into a very soluble product called allantoin) inhibits its activity.
RECONSTITUTION:	Reconstitute with PBS.
ACTIVITY:	Stimulates the caspase-1 activating NALP3 inflammasome, resulting in the production of active IL-1β and IL-18.
SHIPPING:	AMBIENT
LONG TERM STORAGE:	-20°C
HANDLING:	Vortex before use.

Product Images

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Product Description

Specially crystallized and tested on biological activity.

Product Specific Literature References

Gout-associated uric acid crystals activate the NALP3 inflammasome: F. Martinon, et al.; Nature 440, 237 (2006) Abstract

General Information

BACKGROUND/TECHNICAL INFORMATION

Activation of THP-1 cells by uric acid:

1. THP-1 cells are cultured in RPMI 1640 supplemented with 10% heat inactivated fetal bovine serum
    (FCS), 50μM β-mercaptoethanol, 2mM L-glutamine and penicillin/streptomycin. THP-1 cells are grown
    in suspension to a density of 1.5x10⁶ cells/ml in tissue culture flasks.
2. Treat THP-1 cells for 3 hours with 0.5µM PMA (Prod. No. ALX-445-004) the day before stimulation.
3. Wash cells with PBS and plate them in a 12-well plate at a density of 6x10⁵ cells/well in RPMI medium
    (this increases the phagocytic properties of the cells and induces a constitutive production of proIL-1β).
4. The next day, wash cells with PBS and add 500µl Optimem (Invitrogen).
5. Stimulate the cells for 6 hours by addition of 50-200µg/ml of the suspension of uric acid (vortex the
    suspension just before use).
6. As a positive control, 5mM of ATP or 50µg/ml of LPS from E. coli, Serotype R515 (Re) (TLRgrade™)
    (liquid) (Prod. No. ALX-581-007) are incubated for 6 hours.

Detection of the mature IL-1β in the supernatant of uric acid-activated THP-1 cells:

1. Collect the supernatant of each well (500µl). Add 500µl methanol and 125µl chloroform.
2. Vortex and centrifuge for 5 min. at 13’000 rpm.
3. Remove the upper phase.
4. Add 500µl methanol and centrifuge for 5 min. at 13’000 rpm.
5. Remove the supernatant, dry the pellet for 5 min. at 50°C.
6. Add 60µl of 1x loading buffer and DTT (final concentration of 0.1M).
7. Mix and boil before loading on a 15% SDS-gel and transfer onto nitrocellulose membrane. The mature
    IL-1β is detected using the cleaved IL-1β (Asp116) antibody (Cell Signaling).

--> Note: Detection of IL-1β in the supernatant of uric acid-activated THP-1 cells can also be performed by ELISA.
--> Note: Sonication of MSU improves its biological activity in murine cells.

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