MLLT 106 – 
Introduction to Clinical Chemistry: Lesson 2 Review NOTES

Urine Collection

The  type  of  specimen  required  for  a  urine  test  depends  on  the  nature  of  the  test that  is  ordered

EXAMPLE - quantitative  chemical  tests (measurement of urine calcium) , require  a  different  type  of  specimen than is required for a routine urinalysis

• Urine specimens are usually  collected  by  the  patient

        patient must  be given specific instructions about how to collect the urine         specimen

• Urine  specimens  must  sometimes  be  transported  to  a  laboratory  for  testing

• Personnel  must understand  transport requirements and  limitations

        at  the collection  site

        transport  service

        receiving  laboratory

Common  types  of  urine  specimens  submitted  for  laboratory analysis are:

• Random urine specimen

• Fasting or first morning urine specimen

• Clean-catch urine specimen

• Timed specimen (i.e., 24-hour urine specimen)

All urine specimens  should  be collected by the midstream procedure

• except  24-hour  urine  specimens

Midstream urine specimen - the  patient collects only the middle portion of the urine flow

Specimens for Routine Urinalysis

First  morning specimen:

• preferred  specimen  for  routine  urinalysis

• obtained  immediately  on  arising

• normally  more  concentrated  than a random  specimen

• usually has an acid pH - helps preserve any cells present

• can  be  designated as a fasting  specimen - if  collected before the patient has eaten

Random urine specimens:

• most common type of urine sample

• specimens obtained at any time of day

• without regard to food intake

• may not accurately reflect patients' health – may be too diluted causing values to be falsely lowered

Clean-catch  urine  specimen:

• required  when  urine  is  to be  cultured  for  microorganisms

• requires cleansing of the urethral opening before the urine is collected

Timed Specimens: Twenty- Four–Hour Urine Specimens

• Most  quantitative  urine  tests  require  a 24-hour  urine  specimen

• 8-hour  urine  specimens  can be  used  for  certain tests

• Usually  performed  in large hospital or reference laboratories

Collection procedures for  timed  specimens  must  be  followed  carefully  to  ensure that  the  laboratory  has  a  complete  specimen  to  examine

Examples of analytes that can be measured in 24-hour urine specimens - Results are expressed as analyte units per 24h

• Protein

• Creatinine

• Urobilinogen

• Calcium

• Hormones

• Amino acids

• Metabolic  products

• Heavy  metals

• Drugs

Collection of Urine by Catheter

• Occasionally,  urine  for  culture  or  routine  analysis  must  be  collected  by  catheterization

• a  procedure  normally  performed  by nursing  service  personnel

• Catheterization  is  required  when  a patient  is  unable  to  urinate  independently

• A  catheter  is  inserted into  the  bladder  through  the  urethra  and  the  urine  is  collected directly into the appropriate collection container

Suprapubic aspiration collected by inserting needle directly into bladder & aspirating

Pediatric Urine Specimens

• A special  urine  collection  bag  is fitted  to  the  skin  surrounding  the  urethra 

        from  infants  or  small  children who  cannot  urinate  on  demand

• Once  sufficient  urine is  obtained,  the collection bag is removed, and  the  urine  is  transferred into a urine container

Several  sizes  and  shapes  of  containers  are  available  for  collecting urine specimens

Random and first morning specimens: 

• collected in  lidded, disposable  containers

• minimum volume for routine urinalysis is usually 12 mL, but 50 mL is preferable

Clean-catch  specimens:

• urine  containers  must  be  sterile  

24-hour  urine  specimens:

• large,  amber,  opaque  containers

• capable  of holding  at  least 3 to 4 liters (L) 

• often contain preservatives

Labeling Urine Specimens

Urine  specimens  must  be  labeled  clearly:

• patient’s name and identification number

• date and time of collection

Labels must be placed  on  the  container,  not  on  the  lid

Timed specimens - the  start  and  stop  times  should  be  included  on the label or requisition form

LABELLING AND PROCESSING

Proper labeling:

Helps avoid testing delays, which can compromise patient care

Urine specimens should be labeled with the same identifying information as blood specimens

• since many body fluids are similar in appearance, labeling should include the type and/or source of the specimen

Label should be applied to the container, not the lid

• the lid is removed for testing

• to avoid misidentification

SPECIMEN LOG-IN

Time the specimen is received by the testing site should be recorded 

• on the laboratory log,  requisition  form, or specimen label

Check requisition form  - specimen collection meets the criteria for the type of test to be performed

The  specimen  volume  should  be  observed  to  be sure  it  is  sufficient  for  testing

• Usually, a minimum of 15 mL

The  guidelines  in  the  SOP  manual  should  be  followed  when urine volume is less than specified for the test procedure

Urine  specimens  should  be examined within 2 hours of collection

• except  24-hour tests

Deterioration  of  the  specimen  can  be  prevented  by:

• refrigerating the urine at 4° to 6°C for 6-8 hours in the dark in a lidded container

• adding a chemical preservative (not commonly done)

Test results are affected by the way urine specimens are handled  after  collection

Refrigeration of urine can slow bacterial growth - it does not preserve urine sediment components i.e. casts and  cells

Refrigeration protects labile urine components  - ketones, bilirubin, and  urobilinogen

• deteriorate more rapidly at room temperature

• When urine decomposes, the pH becomes more alkaline

Urine rapidly undergoes changes if left at room temperature

Decomposition of urine begins within 30 minutes after collection

Specimens left at room temperature will soon begin to decompose:

• Urea-splitting bacteria produces ammonia

• Ammonia forms ammonium ions = an increase in urine pH  (normal urine pH is 5-7)

• Ammonia like odor is produced

• Increased pH causes decomposition of casts and certain cells

Specimen may be kept 6 to 8 hours under refrigeration with no chemical preservative added

Specimens can be frozen (at 24°C to 16°C) after collection

Common chemical preservatives are hydrochloric acid, boric acid, and acetic acid

Preservatives 

• Prevent  bacterial  growth

• Preserve certain chemical or microscopic components of urine

• Must not interfere with the tests  

• Should  be  used  in  the  correct preservative-to-urine  ratio

Chemical  preservatives can be added to urine  specimens that must be transported

• chemical  preservatives slow the growth of bacteria

• slows the destruction or decomposition of other urine components

Buffered boric acid - added to prevent further growth of any bacteria present 

• When urine  is  to  be  cultured

• Allows urine to be kept at room temperature while still providing results comparable to those of refrigerated urine

Containers  for  24-hour  urine  specimens  usually  have preservatives added before the urine is collected

Preservatives 

• The least hazardous type of preservative should be selected

• Added to the collection container before the urine collection begins

Common 24-hour preservatives:

• Hydrochloric acid

• Boric acid

• Acetic acid

• Toluene

Warning labels should be placed on the container

• A corresponding material safety data sheet should be given to the patient, and the health care provider should explain any potential hazards

• Preserved urine specimens can be stored at room temperature until time of testing

• Product claims regarding the duration of preservative potency should be obtained from the particular manufacturer

Toluene:

• A solution lighter than urine or water

• Prevents the growth of bacteria by excluding contact of urine with air

• A thin layer of toluene is added, just enough to cover the surface of the urine

• The toluene should be skimmed off or the urine pipetted from beneath it when the urine is examined Toluene (toluol) is the best all-around preservative because it does not interfere with the various tests done in the routine urinalysis

Formaldehyde (formalin):

A liquid preservative

Acts by fixing the formed elements in the urinary sediment, including bacteria

•  May interfere with the reduction tests for urine sugar

•  May form a precipitate with urea that interferes with the microscopic    examination of the sediment

Preservative tablets that produce formaldehyde are commercially available

The tablets are more convenient to use than the liquid formalin and do not interfere with the usual chemical and microscopic examination

Thymol:

• A crystalline substance

• Works to prevent the growth of bacteria

• May interfere with tests for urine protein and bilirubin

BD Vacutainer Plus Plastic UA Preservative Tube 

• contains a proprietary additive (chlorhexidine, ethylparaben, sodium propionate)

• maintains sample integrity of up to 72 hours without refrigeration

When a specimen is directly transferred from a collection cup into a preservative tube, it provides a stable environment for the specimen until testing can be conducted and reduces the risk of bacterial overgrowth or specimen decomposition

Urinalysis specimen collection kit (container, transfer device, and preservative transport tube).

Specialized additives: 

• Nitric acid for mercury analysis

• Sodium bicarbonate and ethylenediaminetetraacetic acid for porphyrins

• Sodium bicarbonate for urobilinogen analysis

Boric acid:

Most common preservative of urine for C&S testing is

Comes in tablet, powder, or lyophilized form

• Nonbuffered boric acid may be harmful to certain organisms

• Buffered boric acid preservatives can reduce the harmful effects of the preservative on the organisms

C&S preservatives are designed to maintain the specimen in a state equivalent to refrigeration by deterring the proliferation of organisms that could result in a false-positive culture or bacterial overgrowth

Suspected UTI - urine culture  and  sensitivity  (C&S)  is  ordered

Urine must be collected, processed, and transported in a manner that  prevents  contamination

• If  both culture and routine  urinalysis  are  to  be  performed  on  the  same specimen - the urine must be sent to the microbiology department first for culture, because aseptic techniques are not used in routine urinalysis  procedures

When  urine  to  be  cultured  must  be  transported  over  a  distance - a  kit  containing  a  vacuum  tube  and  sterile straw can  be  used 

• These  kits  work  similar  to vacuum  blood  collecting  systems

• An  aliquot  of  clean-catch  urine is  drawn  into  a  sterile  vacuum  tube  containing  a  preservative  that is not toxic to microorganisms in the specimen

Urine culture and sensitivity (C&S) test kit (Sterile Cup, preservative, urine tube, and castile soap towelettes).

Midstream Collection Method used for:

• First morning

• Random

• Clean-catch specimens

Patient Instructions:

1. Begin  voiding  into  the  toilet

2. Interrupt the  urine  stream

3. Collect  only  the  middle  portion  of  the  urine stream  in  the  specimen  container

This  is  to  prevent  contaminating the specimen with epithelial cells, microorganisms, or mucus from the urethra

Clean-Catch Collection  Method  used  for:

• urine  is  to  be  cultured

Provide the patient with a kit containing towelettes and a sterile urine container

Patient Instructions:

1. Cleanse the urethral opening

2. Carefully collect the urine specimen using the midstream collection method

3. Avoid  touching  the  inside  of  the  sterile container

Written or  pictorial  instructions  that  are  understandable to all patients should be available and should also be posted in the restroom near the toilet

ROUTINE SPECIMENS:

• collected in a clean, dry container

• specimen should be fresh

Routine screening - a freshly voided, random, midstream urine specimen

Other routine urinalysis, including protein content and urinary sediment constituents - concentrated first morning specimen is preferred

CATHETERIZED SPECIMENS:

• Obtained by a physician/nurse - introducing a catheter into the bladder, through the urethra, for the withdrawal of urine

• Required under special circumstances or obtaining a sterile urine specimen for bacteriologic examination

• The risk of introducing infection is always present when an invasive procedure such as catheterization is performed

• Under most conditions, a free-flowing (midstream) voided specimen is satisfactory for bacteriologic cultures

COLLECTION OF TIMED URINE SPECIMENS:

Patient Instructions: 

1. The bladder is emptied at the starting time, such as 8 am, and this time is noted on the collection container

2. The first urine voided at the beginning of the collection is always discarded

3. All subsequent voiding's are collected and put into the container, up to and including the urine voided at 8 am the following day

4. This last urine specimen will complete the 24-hour collection

These timed collection specimens are preserved by refrigeration between collections

• the appropriate chemical preservative added to the container before the beginning of the collection process

The total volume of the timed collection sample is measured and recorded

• the sample well mixed, before a measured aliquot is withdrawn for analysis

The volume  of  a  24-hour urine  specimen  must  be  measured  and  recorded

• the  volume  measurement  is  used  in  calculating  the  test  results

Urine  volume  measurement:

1. use a  large  graduated  cylinder

2. urine  is  returned  to  its  container

3. total  volume  is  recorded on the specimen label and the requisition form

If the 24-hour volume is unexpectedly low, the laboratory can measure  the  creatinine  level  of  the  urine  specimen  and  correlate  it with the urine volume

• At least 1g of creatinine should be excreted in  a  24-hour  period;  a  value  less  than  this  suggests  incomplete collection.

Specimen volume is not  recorded for routine  urinalysis

Reference Ranges for 24-hour Urine Volumes

The volume of urine normally excreted in 24 hours varies according to age:

• Newborns produce between 20 and 350 mL of urine in 24 hours

• By the age of 1 year, 300 to 600 mL/24 hours is  normal

• For  10-year-olds,  the  24-hour  urine  volume  can  range from  750  to  1,500  mL

• Adults  produce  from  750  to  2,000  mL  in 24 hours, with 1200–1500 mL per day being the average

Factors that influence  urine  volume:

• Fluid intake

• Diet

• Time  of  day

• Fluid  lost  in  exhalation  and  perspiration

• Hormone  levels

• Status  of  renal  and  cardiac  functions

Urine  excretion  during  the  day  is  usually  three  times  the volume  excreted  at  night

• Nocturia - excessive production of urine at night

Polyuria - Excessive  production of urine  (over 2000 mL/24  hours)

Oliguria  - Insufficient production of urine  (less  than  500  mL/24  hours)

Anuria - Absence of urine production

Diuresis - Any increase in urine volume, even if temporary
Nocturia - excretion of more than 400 mL urine at night

URINE FOR CULTURE:

When both a bacteriologic culture and a routine urinalysis are needed on the same specimen:

• the culture should always be done first

• then the routine tests

• to avoid contamination of the specimen before culturing on bacteriologic media

URINE FOR CULTURE:

• A clean-catch, midstream urine specimen is desirable for culture

• It is important that the glans penis in the male and the urethral orifice in the female be thoroughly cleaned with a mild antiseptic solution 

Instruction to the patient:

1. Urinate forcibly and allow the initial stream of urine to pass into the toilet or bedpan

2. Throughout the urination process for the female, the labia should be separated so that no contamination results

3. The midstream specimen should be collected in a sterile container, and no portion of the perineum (female) should come in contact with the collection container

4. After the specimen has been collected, the remaining urine is discarded

Urine drug  screens  - can  be  required  in  a  number  of  situations:

• participation in athletic events

• job applications

• cases of suspected  drug  abuse

Random urine specimen  is used  for  drug  screens

Additional documentation is required - guarantee the reliability of the collection procedure and the identity of the person submitting the specimen

Each laboratory that handles specimens for drug screening must follow the written protocol in the  laboratory’s  SOP  manual

Urine drug  screens  

Chain-of-custody  of  the  specimen must  be  documented  to  safeguard  against  possible  tampering  and  to  guarantee  the  specimen’s  integrity 

Some requirements usually included in a protocol for urine drug-screen collection are:

1. Photo identification

2. Signed consent of patient/donor

3. Use  of  special  collection  kits  provided  to  the  patient  by  the laboratory

4. Inspection  of  bathroom  before  and  after  collection,  with  a monitor stationed outside during collection

5. Urine temperature measured and recorded immediately after collection

6. Specimen labeled in presence  of  donor,  sealed  in  outer container, and secured until transported to testing agency

Urine Cytology Studies – looks for the presence of abnormal cells that shed from the urinary tract into the bladder

Performed to detect: 

• Cancer

• Cytomegalovirus

• Other viral and inflammatory diseases of the bladder and other structures of the urinary system

Cells from the lining of the urinary tract are readily shed into the urine

• a smear containing them can easily be prepared from urinary sediment or filtrate

The smear is stained by the Papanicolaou (PAP) method and examined under a microscope for the presence of abnormal cells

A fresh clean-catch specimen is required for the test

• specimen should be examined as soon after collection as possible

• can be preserved by the addition of an equal volume of 50% alcohol

COMMON SPECIMEN REJECTION - URINE

1. Unlabeled or mislabeled samples

2. Requisition does not match specimen request

3. Leaky containers

4. Contaminated samples - For example, a urine sample should not contain stool or paper

• In the case of a contaminated sample, request a new sample

5. Inappropriate sample sources

• Samples that do not conform to the type of sample needed for the requested test(s) are unacceptable 

• For example: 24–hour urine samples are unacceptable for routine bacterial cultures

• If an incorrect or inappropriate sample type is received, request a new sample and specify the proper sample for the test requested

6. Delayed transport time and sample processing

• If the time between sample collection and receipt is too long for a valid test to be performed, with respect to sample requirements for the requested test(s), request a new sample

• If a sample was received after prolonged delay but is not rejected by the laboratory, document it and indicate the length of time after collection that the sample was received

Actions for when samples are rejected

• If the unacceptable sample can be replaced, notify the requesting healthcare provider

• Document the reason for the sample unacceptability and request another sample

• Do not discard the sample until the patient’s healthcare provider has confirmed that another can be collected

• If the patient has already been started on antimicrobial therapy or if a repeat sample cannot be collected, this must be documented

• If a repeat sample is not available, document the problem and proceed with the test if possible

Urine specimens can be disposed of in ordinary sewage systems, such as laboratory sinks

followed by flushing with water and use of disinfectant

Urinalysis requires a quality assessment program to ensure that results of testing are meaningful

The preanalytical stage of urine testing consists of the following six phases:

1. Test ordering (requisition)

2. Sample collection

3. Specimen transport to the laboratory

4. Specimen receipt in the laboratory

5. Preparation of samples for testing

6. Transportation of samples to the section of the laboratory where testing occurs

Common areas of preanalytical variability in urine testing include:

• Patient-related factors

• Specimen collection

• Specimen identification

• Specimen labeling

• Specimen transfer

• Specimen transport

• Specimen processing