bSLSBio LifeSpan BioSciences, Inc. Protein Creatinine Ratio (PCR) Assay Kit (Colorimetric) LS-K318-100 (100 Tests) • Store at 4°C Introduction PROTEIN is filtered out of urine by the glomeruli of the kidneys. Albumin is the most common serum protein, thus the majority of the protein in urine is albumin. A damaged kidney will allow some protein through into the urine, the less protein in urine the better. Elevated protein levels in urine is called microalbuminuria or proteinuria, which typically arises due to type 1 diabetes, type 2 diabetes, or high blood pressure. CREATININE is synthesized in the body at a fairly constant rate from creatine. In healthy individuals, creatinine secretion is independent of diet and is fairly constant. The creatinine clearance test has become one of the most sensitive tests for measuring glomerular filtration rate. PROTEIN/CREATININE RATIO (PCR) remains the simplest and most convenient test for proteinuria. Other Methods such as 24 hour urine test or timed urine test require strict adherence to sample collection protocol. Since the protein concentration is normalized to creatinine secretion, the urine sample can be taken at any time and no diet or liquid restrictions are necessary for sample collection. Key Features • Sensitive and accurate. Use 20 pL samples. Linear detection range in 96-well plate: 1 - 20 mg/dL Protein and 1 -150 mg/dL Creatinine. • Fast and convenient. No sample pre-treatment is needed. Simple 10- minute "add-incubate-read" procedure. • High-throughput adaptable. The procedure can be readily automated for processing thousands of samples per day. Applications • Direct Assays: Protein creatinine ratio determination in urine samples (rat, mouse, human, not species specific). • Drug Discovery/Pharmacology: effects of drugs on protein and creatinine concentration, metabolism, and excretion. Materials Not Supplied Pipetting devices, centrifuge tubes, clear flat-bottom 96-well plates (e.g. VWR cat# 82050-760), and plate or cuvette reader. Storage The kit is shipped at room temperature. Store all reagents at 2-8°C. Shelf life: 12 months after receipt. FOR RESEARCH USE ONLY! Not for use in humans. LifeSpan BioSciences, Inc. • 2401 Fourth Avenue, Suite 900, Seattle, WA 98121 www.LSBio.com • (206) 464-1554 • [email protected]

Protein Creatinine Ratio (PCR) Assay Kit (Colorimetric) LS-K318-100 (100 Tests) • Store at 4°C Assay Procedure Samples can be analyzed immediately after collection, or stored in aliquots at 4°C or –20°C for 7 days. Avoid repeated freeze-thaw cycles. If particulates are present, centrifuge sample and use the clear supernatant for the assay. Equilibrate all components to room temperature. Protein Determination 1. Samples are run in duplicate. Transfer 20 μL of each sample into four separate wells: two Sample wells and two Internal Standard wells. Add 5 μL dH2O to Sample wells, and 5 μL of Standard...

bSLSBio LifeSpan BioSciences, Inc. Protein Creatinine Ratio (PCR) Assay Kit (Colorimetric) LS-K318-100 (100 Tests) • Store at 4°C Protein concentration of a Sample is calculated as [Protein] = Pimple - ODb^nk x 10000><n ( /dL) 1 J ODstandard - ODsample Creatinine concentration of a Sample is calculated as ODsample- ODblank ODstandard - ODsample Protein Creatinine Ratio of a Sample is calculated as [Pr°te'n^ (vg/mg) [Creatinine] Protein Creatinine Ratio where ODsample, ODstandard, and ODblank are the optical density values of the Sample, Internal Standard, and Blank wells, respectively. 10,000...