Urine samples

COLLECTION AND STORAGE

 

Urine should be collected and stored in test tubes as provided (Urine Monovette, Sarstedt, Germany; article number 10.253.001).  The recommended test tubes contain 15 mg/mL boric acid to enhance preservation.

Approximately 10 mL should be collected.  Optimal is the second morning urine, but any urine sample collected between 7 AM and 11 AM works fine. The first morning urine is less suited, as it shows higher variability in peptide content, probably due to differences in urinary output during the night. Urine samples obtained in afternoon and evening also show a higher degree of variability, and generally lower concentration of peptides, which may result in the sample being of too low quality to be properly evaluated.  For spot urines, mid- stream urine should be collected.  Taking an aliquot from 24‑h urine samples is also possible, as long as the 24‑h urine sample is collected in the presence of NaN3 (10 mL of a 5% solution) as preservative and hence does not contain microbial contamination.  In general, samples should not be turbid and should be frozen without any further manipulation.

To assess the urinary proteomic profile (UPP) requires an 8-mL urine sample. The UPP does not change when urine is stored for 8 hours at room temperature and for 3 days at 4 °C, thereby providing a wide time window for handing samples.  The urine can be indefinitely stored at ‑20 °C without UPP alteration (Proteomics Clin Appl 2010; 4: 464-74 // Clin Biochem 2013; 46: 432-43).  Optionally, if possible, the urine samples may be stored at ‑40 °C or at ‑80 °C a few days before shipment to Hannover, Germany, because at this low temperature, thawing of the urine takes more time, thereby facilitating the integrity of the UPP during shipment. 

 

THE URINARY PROTEOMIC PROFILE

Urine contains over 21,000 peptide fragments, of which 5070 have been sequenced and allow to identify the parental protein.  These data will be made available to the UPRIGHT-HTM research consortium.  However, for clinical purposes, patients and their caregivers will be given information on the urinary levels of CKD273 and HF1.   The multidimensional urinary classifier CKD273 (consisting of 273 peptides) predicts progression of CKD earlier than micro-albuminuria does (KI Reports 2017; 2: 1066-75).  HF1 (85 peptides) is a marker of subclinical left ventricular dysfunction (Proteomics Clin Appl 2019; 13: 1800174).  Both CKD273 and HF1 have been extensively validated in longitudinal patient and population studies.  The optimised diagnostic (actionable) thresholds are 0.154 for CKD273 and -0.350 for HF1.     

The number of urine samples processed for UPP amounts to: 105 for the Nigerian centres, 27 for Kraków, Poland, and 7 for Ljubljana, Slovenia.

 

 

 

For more information, do visit the web pages of Mosaiques-Diagnostiques via https://mosaiques-diagnostics.de/mosaiques-diagnostics/. 

 

SHIPMENT OF URINE SAMPLES

If you would like to ship urine samples to Mosaiques-Diagnostics GmbH, Hannover Germany (team of Professor Harald Mischak), please, contact the study coordination team via jan.staessen@appremed.org or dewei.an@appremed.org, so that the coordination centre can liaise with DHL.

Urine Monovette When you are preparing a shipment, please, take care of the following important points:

  1. Make sure that the caps of the test tubes are tightly closed to prevent leakage of urine.

  2. Each Urine Monovette should be labelled with a unique UPRIGHT-HTM identification number, using waterproof ink.  
  3. Do check the screening and pre-randomization eCRFs, to make sure that all information is duly filled out.  It is indeed meaningless to engage in complex urinary proteomic profiling if the clinical and biochemical data are incomplete or incorrect.

  4.  Do mention in your request, the full institutional address (postal address, including zip [postal] code), the person responsible for being contacted by DHL (full first and alst names, email address, and telephone number including a cell phone number).  The cell phone should be enabled to receiving short text messages.

  5. Make sure that there is an electronic inventory in Excel format with the following information: the center number, the unique UPRIGHT-HTM identification number, the bag number (see point 7), sex, date of birth, and date of the urine collection.  The electronic inventory should be printed and added as a paper document in a plastic sheet inside the shipment box.  In addition, the electronic inventory should be emailed a few days prior to the intended shipment date to Prof Harald Mischak (mischak@mosaiques.de) and the coordinating team at APPREMED (dewei.an@appremed.org; yuling.yu@kuleuven.be; jan.staessen@appremed.org).

  6. For packaging, do use an insulating Styrofoam box.  The coordinating office will organise a delivery to Hannover, Germany, within 48 hours after the sample box is collected at your centre.  If the latter is feasible, there is no need for dry ice or to add grozen gel bags.  In the unlikely case that the service might take more than 48 hours including frozen gel bags in the package is useful to preserve urine samples.  We suggest removing the urine samples from the deep freezer only 60 minutes before the package is collected.  

  7. APPREMED has a license for shipping the urine samples (biological samples) as "dangerous goods" to be coded as UN3373.  This requires that the package should comply with the instructions for UN3373 shipments.  Two issues are important.  First, the package should be externally labeled with the UN3373 code.  Second, to prevent urine leakage from the test tubes, there should be a "double seal".  Combine 5-10 test tubes in one plastic bag (first seal).  Each bag should be numbered.  A plastic bag enclosing all small bags can function as the second seal.  Detailed packaging instructions for UN3373 specimens are available at https://www.purolator.com/en/services/purolator-specialized-services/dangerous-goods/un3373-biological-substance.  

  8. The additional information needed consists of the number of urine samples, the weight of the package in kilograms, and the volumetric weight of the package to be calculated as height × width × depth in centimeter.  Note that for prizing by DHL the volumetric weight takes precedence over the true weight.  Therefore, use the smallest box possible.   To be cost-effective, with some exceptions, shipments should not include less than 25 samples, unless the time delay from randomisation of any patient would exceed ~6 months.  

  9. In your request, do mention the preferred date/time of the sample collection.

  10. Once the shipment has been arranged, the coordinating office will email the waybill to the contact person at the clinical centre.  The waybill should be given to the person collecting the shipment.  

  11. The person responsible for making the shipment available to DHL at the point of departure, in most cases, will receive emails from the local DHL agency to confirm the date and time of the collection of the shipment.  These emails must be answered immediately.