Aug 08, 2019
Version 2
U Michigan - Spot urine collection V.2
- Jeff Hodgin1
- 1University of Michigan - Ann Arbor
- Mouse Metabolic Phenotyping CentersTech. support email: info@mmpc.org
Protocol Citation: Jeff Hodgin 2019. U Michigan - Spot urine collection. protocols.io https://dx.doi.org/10.17504/protocols.io.56fg9bn
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: August 06, 2019
Last Modified: August 08, 2019
Protocol Integer ID: 26535
Keywords: urine sample, albumin/creatinine ratio, glomerular nephropathy
Abstract
Summary:
Collection of urine from experimental animals is a basic requirement to detect albumin/creatinine ratio for glomerular nephropathy. This is a non-invasive, simple, quick, and inexpensive method for collecting spot urine from mouse when a small volume of urine (10-500ul) sample is required.
Materials
MATERIALS
Customized spot urine cage- a clear plastic container for lean or obese mouse. 6 ½” x 4 7/8” x 3 7/8” for lean mouse. (7 1/8” x 6 1/2” x 4 ½” for obese mouse. AmazonCatalog #SRERILITE 177718612 Flip Top Sto
SMALL PARTS INC. Stainless Steel 304 Mesh #10 0.023Wire Mesh Cloth Screen 12’’x24’’ Amazon
RAFTSMAN 3/8 INCH Electric Drill and 1/8 INCH Drill Bit Sears
Metal Cutter and Flyer from Home Depot
Paper towels disinfectant spray anti-bacterial soap cleaning brush and sponge Fisher ScientificCatalog #Supplied by Lab. Animal unit
Reagent Preparation:
Reagents and Materials:
Procedure: Careful during working with Metal cutter, electric Drill, Stainless Steel meshes.
Use lab personnel protections e.g. Safety glasses, gloves, lab coat
Create 6 evenly spaced holes in the lid of the container – 3 on top and 3 at bottom using an electric drill using 1/8” drill bit. Also drill 2 holes each on four sides of the container. Using a metal cutter cut out a 14.5 cm by 9.5 cm piece of stainless steel wire mesh to create a base for mouse that will fit inside the lean mouse cage. Then bend all four sides of the wire mesh using a flyer to create 1 cm perpendicular bent on each sides. Insert the wire mesh base inside the container with the side bents pointing downward. The base should have a snug-fit inside the container about 1” above the bottom. Create similar way for obese mouse cage. Wire mesh need to be washed and autoclaved before using.
It is important to acclimatize the mouse in the metabolic cage for about half hour to one hour every day for 3 consecutive days before starting urine collection.
Prepare appropriate number of urine collecting tubes by labeling a tube with identification number for each animal.
To maintain SPF housing, place cages in an operational laminar flood hood.
Place the appropriate mouse in the metabolic cage and replace the cage lid. Be certain the lid is secure. Put one mouse per box and label the top of the box with the mouse number.
Place the mouse cage card in front of metabolic cage. Make sure mouse cage card has all the necessary information such as mouse identification number, mouse protocol number, emergency contact number, Principal Investigator name, date, and time period of 4 hour urine collection, also “DO NOT FEED” sticker (Provided by Lab Animal Facility)
Because mice need to be left without food or water for 4 hours, an exception must be completed.
Collect urine for 4hour period without food and water access.
Check on mice every hour to monitor mouse health conditions.
After 4hour remove the mouse from the metabolic cage and replace in the original housing cage and carefully collect the urine.
Collect urine using a pipette (P200 is usually best) and place in a 1.5 mL micro centrifuge tube.
Spin urine at 2000 rpm for 5 min. Remove supernatant and place in a clean 1.5 mL tube.
Store urine samples at 4ºC until ready for assay in short period and at -20ºC or -80ºC for a longer period.
Clean metabolic cage with appropriate disinfectants. Hot, soapy water may be used on the acrylic parts. All cage parts should be sprayed with a bactericidal, virucidal, and fungicidal (e.g., Clidox® or Spor-Klenz ®) and wiped down. Dry each part and reassemble