Dried Urine Analysis: Improved Collection, Shipment, Processing, Testing and Storage of Samples

by Theodore Zava

Introduction

Dried urine collection and testing from laboratory-grade filter paper is a relatively new process that is increasing in popularity. Similar to blood spot collection, in which blood drops are collected on a filter card and allowed to dry, the collection procedure for dried urine is simple and can be completed at any location. ZRT Laboratory first realized the feasibility and benefits of collecting and testing dried urine for elements with the successful development and validation of urinary iodine and creatinine tests.¹ These tests were then followed by the successful development and validation of a multielement dried urine assay including iodine, bromine, selenium, arsenic, cadmium, and mercury, which will be used in this article to help describe the benefits of dried urine testing.

Volume of Urine Required is Minimal

Depending on the size of the collection device, it takes very little urine to saturate a filter strip (2 mL for a 2.5 × 6 cm strip of Whatman Grade 903 Specimen Collection Paper). Unlike blood or saliva, urine is plentiful and can be collected throughout the day. Even without urinary urge, sufficient urine can usually be collected to saturate a filter strip.

Collection, Storage, and Shipment of Urine on Filter Strips is Simple

Collection of urine onto filter strips is very simple and can be completed in places such as at home, a doctor’s office, or in remote locations. Urine is applied to a laboratory-grade filter paper either by dipping the filter paper in urine collected in a vessel or by urinating directly on the strip. The urine quickly saturates the filter strip and can be removed from the urine source immediately after it is applied. After at least 4 hours of drying, the sample is ready to be stored or shipped to the testing laboratory for analysis.

Stability of Elements in Dried Urine Is at Least

1

Month

Analyte stability under different environmental conditions is a major concern for liquid samples, as most are transported and stored prior to testing. Without the addition of antimicrobial/stabilizing agents or refrigeration/freezing of liquid samples, analytical results may be compromised. Liquid urine mercury analysis requires extra care during collection, transportation, and storage of samples to prevent analyte loss, often requiring a preservative such as hydrochloric or sulfamic acid to prevent the adsorption or transpiration of mercury in collection vessels.² Addition of these stabilizers can dilute or contaminate the sample, or interfere directly with analysis.³ Without the use of preservatives, urine mercury may be unstable, especially at higher temperatures or lower concentrations.^3,4 Because of mercury’s instability in liquid samples, testing laboratories often recommend or require refrigerated or frozen transportation of urine for mercury analysis, which is expensive and not always an available option. Stability tests show that mercury analyzed from human urine dried on filter paper is stable at room temperature, refrigerated, or frozen for at least 1 month (Figures 1–3).

Figure 1: Mercury is stable in dried urine samples stored at room temperature over 27 days.

Figure 2: Mercury is stable in dried urine samples stored at 4

ºC over 27 days.

Figure 3: Mercury is stable in dried urine samples stored at

−20

ºC over 27 days.

Transport of Dried Urine Has Few Restrictions and Is More Economical

Dried urine filter strips are very small and weigh very little in comparison with liquid urine collections. Multiple urine filter strips can be packed inside a small shipping device with plenty of room to spare (Figure 4). Since refrigerated or frozen transport is not necessary, shipment around holidays or late in the week (involving extended transfer times) is not an issue.

Figure 4. Dried Urine Filter Paper Transportation and Storage

Bacterial Growth is Not an Issue with Dried Urine

A major issue in liquid urine samples that aren’t frozen is bacterial growth occurring either before, during, or after transportation. Urine dried on filter paper is not subject to rapid bacterial growth as long as samples are properly dried before storage.

Laboratory Preparation for Testing Is Much Easier with Dried Urine

Liquid samples may require sample pretreatment; for example, centrifugation and/or manual transfer of samples (with its inherent risk of spilling) that can significantly increase sample analysis time. Dried urine samples can easily be processed for testing using a laboratory hole-punch into 96-well blocks, reducing sample preparation and analysis time, which translates to a reduced turnaround time for the client.

Accuracy of Element Testing Is Excellent in Dried Urine Specimens

Analyte results from urine dried on filter paper are equivalent to, and for certain tests outperform, liquid urine samples. Liquid external controls with certified analyte concentrations are dried on filter paper, analyzed, and compared with their known values (Table 1). We further check accuracy by running liquid and dried urine in parallel using inductively coupled plasma mass spectrometry (ICP-MS) and plotting results against each other (Figure 5). Some external laboratory controls, once reconstituted from their lyophilized form, are only stable for a few hours, but drying them on filter stabilizes them for over a month.

Table 1: Analyzed Dried Urine Reference Controls Compared

with

Their Expected Values

Figure 5: Dried Versus Liquid Urine Iodine Comparison (n

=

100)

Creatinine Correction Is an Important Part of Dried Urine Testing for Analytes

Creatinine is excreted by the body at a relatively constant rate as a breakdown product of creatine phosphate. It is commonly tested along with other analytes in urine to correct for hydration status.⁵ Without creatinine correction, individual results will vary widely depending on daily fluid intake. Dilute urine generally appears clear, while concentrated urine might be bright yellow. Visually, it might not be possible to determine if filter paper is saturated or partially saturated with urine, making creatinine a useful indicator of proper collection as well as the degree of concentration of the urine.

Summary

Improvements in laboratory technology, such as increased sensitivity and specificity for mass spectrometers, have opened new doors for microsampling and analysis of dried samples. Collection of liquid samples on filter paper have gained popularity over the last decade, led primarily by dried blood spot testing and recent advances in dried plasma and dried urine testing.⁶ Urine dried on filter paper is advantageous for both clinical testing and research studies due to increased analyte stability and small storage requirements, while simplifying sample collection and transport. Although it will take time for dried urine testing to become a part of mainstream laboratory testing, the benefits of dried samples are becoming well known and are being applied to resolve problems associated with liquid sample types.

Notes

1. Zava TT, Kapur S, Zava DT. Iodine and creatinine testing in urine dried on filter paper. Anal Chim Acta. 2013 Feb 18;764:64–69.

2. Zava TT, Zava DT. Dried urine mercury stability and quantification using a direct mercury analyzer. Poster presented at: 12th International Conference on Mercury as a Global Pollutant; June 14–19, 2015; Jeju, Korea.

3. Centers for Disease Control and Prevention (CDC). 2011–2012. Iodine and Mercury Laboratory Procedure Manual. Inductively Coupled Plasma Dynamic Reaction Cell Mass Spectrometry Method 3002.1.

4. Gaines P. Mercury chemical stability [Web page]. Inorganic Ventures. 20 February 2015. http://www.inorganicventures.com/mercury-chemical-stability.

5. Barr DB, Wilder LC, Caudill SP, Gonzalez AJ, Needham LL, Pirkle JL. Urinary creatinine concentrations in the U.S. population: implications for urinary biologic monitoring measurements. Environ Health Perspect. 2005 Feb;113(2):192–200.

6. Déglon J, Leuthold LA, Thomas A. Potential missing steps for a wide use of dried matrix spots in biomedical analysis. Bioanalysis. 2015 Sep 17.

Theodore Zava received his bachelor’s degree in biology from Oregon State University in 2009. He is a research associate at ZRT Laboratory in Beaverton, Oregon, where he developed a test to measure iodine, bromine, selenium, arsenic, cadmium, mercury, and creatinine levels in dried urine. His current research focuses on new test development using dried samples while linking intake and excretion of essential elements and toxic metals to cancer and disease.