A Protocol to Perform Systemic Lipopolysacharide (LPS) Challenge in Rats

Protocolo para realizar reto sistémico con lipopolisacárido (LPS) en ratas

Authorship SCIMAGO INSTITUTIONS RANKINGS

Abstract

Lipopolysaccharide (LPS) is a component of the outer membrane of Gram-negative bacteria. In animals, intraperitoneal administration of LPS, stimulates innate immunity and the production of proinflammatory cytokines. LPS provides an inflammatory stimulus that activates the neuroimmune and neuroendocrine systems resulting in a set of responses termed sickness behavior. The purpose of this protocol is to describe step-by-step the preparation and procedure of application of intraperitoneal injection of LPS in rats, as a guide for those researchers that want to use this assay to mount an inflammatory response. LPS intraperitoneal challenge in rats has been widely used to evaluate antiinflammatory reagents and to address basic scientific questions.

Keywords: Lipopolysaccharide; LPS challenge; LPS stimulation; Endotoxin; Inflammation

Optimal		Minimum
Person	Tasks	Person
A	Weighing of the animals to be injected, preparation of saline solution and LPS solution	A
B	Transport of rats from the vivarium to the laboratory or room where the injection is made and returned to the vivarium
C	Intraperitoneal injection of the animal
3	Total people required according to condition: 99. ←Optimal Minimum→	1

Value A	Value B	Value C	Value D
Total mass of animals (kg)	Minimum mass of LPS required (mg)	Mass of LPS to be measured (mg)	Volume of saline solution to dissolve the LPS (mL)
	[0.5 mg/kg * Value A]=	[Value B* 1.75]=	[Value C / 1 mg/mL] =

169. 170.

Critical Points	Why is it a critical point?	Related care
Storage of the LPS solution	Once prepared, the LPS solution can only be stored for a certain time and under certain conditions to ensure its integrity at the time of injection.	To ensure the stability of the LPS solution, use the following storage conditions, depending on the supplier of the reagent (Sigma): 161. Cooling (2-8 ° C): stability up to 1 month. 162. Freezing (-20 ° C): stability up to 2 years (store in aliquots to avoid repeated freeze / thaw cycles).
Rat injection	When injecting the LPS solution, care must be taken so that the needle does not move inside the abdomen, to avoid lacerating surrounding organs or blood vessels.	Avoid causing harm to the animal: 167. Carrying out a good immobilization of it. 168. Making sure to puncture with the needle at the anatomical site indicated.
-	Puncture of these sites should be avoided, however, if any of these substances are present, it is better to remove the needle from the animal, evaluate the puncture wound, evaluate if the animal requires some treatment and, if feasible, redo the puncture. If the syringe got contaminated, discard the needle and syringe into the sharps bin and biohazard waste bin, respectively, and use a new syringe and a new needle in the next injection.
After the intraperitoneal injection	The animal should be monitored immediately after the injection, about 10 minutes later and the next day. • If bleeding occurs, place gauze and apply pressure. Once bleeding stops, clean the site with gauze and water. 177. • In case of peritonitis, laceration of internal organs and / or infection, consult a veterinarian to assess if the animal can continue in the experiment.

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None Ciudad Universitaria Rodrigo Facio, San Pedro, San José, San José, CR, 2060, 2511-8102, 2234-9207 - E-mail: odontos.fo@ucr.ac.cr

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