With the aid of routine examinations early symptoms of the following four groups can be identified: • Diseases of the kidneys and the urinary tract • Carbohydrate metabolism disorders (diabetes mellitus) • Liver diseases and haemolytic disorders • Urinary infections
Urinary tract Screening parameters: Many renal and urinary tract diseases may be
asymptomatic for a long period of time. Routine urinalysis is recommended as a basic yet fundamental step in identifying
renal damage and/or
urinary tract disease at an early stage, especially in high-risk populations such as
diabetics, the
hypertensive,
African Americans,
Polynesians, and those with a
family history. Specific kidney and urinary tract diseases that can be identified include:
chronic kidney disease,
glomerulonephritis,
proteinuria and
haematuria.
Protein testing Of the routine chemical tests performed on urine, the most indicative of renal disease is the protein determination. Proteinuria is often associated with early renal disease, making the urinary protein test an important part of any physical examination. Normal urine contains very little protein, usually less than 100–300 mg/L or 100 mg per 24 hours is excreted. This protein consists primarily of low-molecular-weight serum proteins that have been filtered by the glomerulus and proteins produced in the genitourinary tract. Due to its low molecular weight, albumin is the major serum protein found in the plasma, the normal urinary albumin content is low because the majority of albumin presented in the glomerulus is not filtered, and much of the filtered albumin is reabsorbed by the tubules. Other proteins include small amounts of serum and tubular microglobulins. Uromodulin produced by the renal tubular epithelial cells and proteins from prostatic, seminal, and vaginal secretions. Uromodulin is routinely produced in the distal convoluted tube, and forms the matrix of casts. Traditional reagent strip testing for protein uses the principle of the protein error of indicators to produce a visible colorimetric reaction. Contrary to the general belief that indicators produce specific colours in response to particular pH levels, certain indicators change colour in the presence of protein even though the pH of the medium remains constant. This is so because protein accepts hydrogen ions from the indicator. The test is more sensitive to albumin because albumin contains more amino groups to accept the hydrogen ions than other proteins. Depending on the manufacturer, the protein area of the strip contains different chemicals. Multistix contains tetrabromophenol blue and Chemstrip contains 3’,3”,5’,5”-tetrachlorophenol, 3,4,5,6-tetrabromosulfonphthalein. Both contain an acid buffer to maintain the pH at a constant level. At a pH level of 3, both indicators appear yellow in the absence of protein. However, as the protein concentration increases, the colour progresses through various shades of green and finally to blue. Readings are reported in terms of negative, trace, 1+, 2+, 3+ and 4+ or the semi-quantitative values of 30, 100, 300 or 2000 mg/dL corresponding to each colour change. Trace values are considered to be less than 30 mg/dL. Interpretation of trace readings can be difficult.
Indicator-H+(Yellow) + Protein → Indicator(Blue-green) + Protein-H+ The major source of error with reagent strips occurs with highly buffered alkaline urine that overrides the acid buffer system, producing a rise in pH and a colour change unrelated to protein concentration. Likewise, a technical error of allowing the reagent pad to remain in contact with the urine for a prolonged period may remove the buffer. False-positive readings are obtained when the reaction does not take place under acidic conditions. Highly pigmented urine and contamination of the container with quaternary ammonium compounds, detergents and antiseptics also cause false-positive readings. A false-positive trace reading may occur in specimens with a high specific gravity.
Hemoglobin and myoglobin testing is clearly visible, it is rare to find examples in such a well conserved condition. The presence of
blood in the urine is, of all the parameters normally tested, the one that is most closely related with traumatic damage to the kidneys or the genitourinary tract. The most common causes of hematuria are:
nephrolithiasis,
glomerular disease,
tumours,
pyelonephritis, exposure to
nephrotoxins, and treatment with
anticoagulants. Non-pathological hematuria can be observed after strenuous exercise and during
menstruation. The normal number of red blood cells in urine should not usually exceed 3 per high power field. A urine test strip showing positive for blood can also indicate
hemoglobinuria, which is not detectable using a microscope due to the lysis of red blood cells in the urinary tract (particularly in alkaline or dilute urine), or
intravascular hemolysis. Under normal conditions the formation of
haptoglobin-hemoglobin complexes prevents glomerular filtration, but if the hemolysis is extensive haptoglobin's uptake capacity is exceeded and hemoglobin can appear in urine. Hemoglobinuria can be caused by hemolytic anaemia, blood transfusions, extensive
burns, the bite of the
recluse spider (Loxosceles), infections and strenuous exercise. The urine test strip test for blood is based on hemoglobin's pseudo peroxidase activity in catalysing a reaction between hydrogen peroxide and the chromogen tetramethylbenzidine in order to produce a dark blue oxidation product. •
Na2[Fe(CN)5NO] + CH3COCH2COOH + 2Na(OH) → Na4[Fe(CN)5-N=CHCOCH2COOH](magenta) + H2O •
Sodium nitroprusside + Acetoacetic acid + Alkali medium → Pink-magenta complex + Water The test does not measure beta-hydroxybutyric acid and it is only weakly sensitive to acetone when
glycine is added to the reaction. However, as these compounds are derived from the acetoacetic acid their existence can be assumed and a separate test is not therefore necessary. Those medicines that contain sulfhydryl groups, such as mercaptoethane sulphonate Na (
Mesna) and
captopril and
L-DOPA can give atypical colouring. A false negative can occur in samples that have not been adequately stored due to volatilization and bacterial degradation.
Liver and blood disorders In many liver diseases the patients often show signs of pathology only at a late stage. Early diagnosis allows appropriate therapeutic measures to be instituted in good time, avoiding consequential damage and further infections. Specific liver diseases and haemolytic disorders able to be identified include
liver disease, (accompanied by
jaundice),
cirrhosis, urobilinogenuria and
bilirubinuria.
Bilirubin test Bilirubin is a highly pigmented compound that is a by-product of haemoglobin degradation. The haemoglobin that is released after the
mononuclear phagocyte system (located in the
liver and
spleen) withdraws old red blood cells from circulation is degraded into its components;
iron,
protoporphyrin and
protein. The system's cells convert the protoporphyrin into
unconjugated bilirubin that passes through the circulatory system bound to protein, particularly albumin. The kidney is unable to filter out this bilirubin as it is bound to protein, however, it is conjugated with
glucuronic acid in the liver to form water-soluble conjugated bilirubin. This conjugated bilirubin does not normally appear in the urine as it is excreted directly from the intestine in
bile. Intestinal bacteria reduce the bilirubin to
urobilinogen, which is later oxidised and either excreted with the faeces as
stercobilin or in the urine as
urobilin. Conjugated bilirubin appears in urine when the normal degradation cycle is altered due to the obstruction of the biliary ducts or when the kidney's functional integrity is damaged. This allows the escape of conjugated bilirubin into the circulation as occurs in
hepatitis and
hepatic cirrhosis). The detection of urinary bilirubin is an early indication of liver disease and its presence or absence can be used to determine the causes of clinical
jaundice. The jaundice produced by the accelerated destruction of red blood cells does not produce bilirubinuria, as the high serum bilirubin is found in the unconjugated form and the kidneys are unable to excrete it. The test strips use a
diazotization reaction in order to detect bilirubin. The bilirubin combines with a
diazonium salt (2,4-dichloroaniline or 2,6-dichlorobenzene-diazonium-tetrafluoroborate) in an acid medium to produce an
azo dye with colouration that varies from pink to violet: Any deterioration in liver function reduces its ability to process the recirculated urobilinogen. :* (1) Reaction on Multistix (in acid medium)
Urobilinogen + p-dimethylaminobenzaldehide → Red dye :* (2) Reaction on Chemstrip (in acid medium)
Urobilinogen + 4-methoxibenzene-diazonium-tetrafluoroborate → Red azo dye A number of substances interfere with the Ehrlich reaction on the Multistix strip: porphobilinogen, indican, p-amino salicylic acid, sulphonamide, methyldopa, procaine and
chlorpromazine. The test should be carried out at room temperature as the reaction's sensitivity increases with temperature. Poorly stored samples can yield false negative results as the urobilinogen suffers photo oxidation to urobilin that does not react. The
formaldehyde used as a preservative produces false negatives in both reactions. The test is a rapid screen for possible infections by enteric bacteria, but it does not replace the
urinalysis tests nor microscopic examination as diagnostic tools, nor subsequent monitoring as many other microorganisms that do not reduce nitrate (
gram positive bacteria and yeasts) can also cause urinary infections. The reactive strips detect nitrite by using the
Griess reaction in which the nitrite reacts in an acid medium with an
aromatic amine (para-arsanilic acid or sulphanilamide) in order to form a
diazonium salt that in turn reacts with tetrahydrobenzoquinoline to produce a pink azo dye. Neutrophil granulocytes are the leukocytes most commonly associated with urinary infections. A positive test for leukocyte esterase normally indicates the presence of bacteria and a positive nitrite test (although it is not always the case). Infections caused by
Trichomonas,
Chlamydia and yeasts produce
leukocyturia without bacteriuria. The inflammation of the renal tissues (
interstitial nephritis) can produce leukocyturia, in particular toxic interstitial nephritis with predominant eosinophils. == Detection limit ==