Interference indices

 
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The sample indexes of lipemia, hemolysis and icterus are part of our chemistry results. They are provided automatically with our analyzer with any chemistry test and are very useful.  These indexes are more objective and consistent than visual assessment of a sample.  Hemolysis from ruptured red blood cells, whether in vitro or in vivo), lipemia (due to increased chylomicrons and/or very low density lipoproteins) and icterus (from hyperbilirubinemia) in a sample can, in some cases, interfere with accurate measurement of clinical pathologic analytes through various mechanisms (for more, see interferences under test basics).

Guides as to which tests are affected at different levels for each index are provided with the chemistry results from Cornell University. These are guidelines only (many are based on data from humans, particularly the product information sheet from the manufacturer, and may not be applicable to animals) and should be used carefully to assess the impact of these preanalytical variables (lipemia, hemolysis, icterus) on the chemistry results. When doing this, it is very important to interpret these values along with the clinical signs displayed by the patient and the remaining laboratory results, i.e. they should never be used alone. Remember that the effect of these interferences on chemistry results are also dependent on the analyzer and methods used. They cannot be transferred from one laboratory’s results to another. These guidelines are given below for each interference with the analyzer used in our laboratory (currently the Cobas 501).

Lipemia index

The number reported under lipemia on our chemistry reports is a measure of the lipid content of the plasma. Index values are obtained by adding increasing amounts of an emulsion of fat to a sample (Intralipid®) and assessing the effect the emulsion has on chemistry values. In reality, the lipemia index correlates weakly to the triglyceride concentration in clinical sample. In our experience, the units for lipemia index correlate to gross estimates of lipemia as follows:

 

Guide to Lipemia Index
Lipemia index Gross appearance
<30 No lipemia
30-60 slightly lipemia (hazy)
60-120 moderately lipemia (milky)
>120 markedly lipemia (creamy)

 

The guidelines below indicate how lipemia, based on the lipemia index, may affect results on our current chemistry analyzer. These guidelines are derived from manufacturer’s recommendations and are based on data from spiking human samples with Intralipid®. These values may not apply to samples from animals and there may be species differences. These guidelines should always be used in conjunction with clinical signs and knowledge of the underlying disease processes as guides for interpreting laboratory data and determining the accuracy of reported laboratory results.

Effect of lipemia (based on lipemia index) on chemistry results (guidelines)
Lipemia index Possible effect on tests
>60 Unknown effect with 5th generation reagents:  Bile acids
≥100 Falsely increased:  Direct bilirubin.
Unknown effect with 5th generation reagents:  Bile acids.
≥150 Falsely increased:  Direct bilirubin.
Falsely decreased
: sodium, potassium (less affected than sodium and chloride), chloride (if triglyceride concentrations are high, e.g. > 600 mg/dL).
Unknown if increased or decreased: ALT, AST, bile acids.
≥300 Falsely increased:  Direct bilirubin, TIBC.
Falsely decreased
: sodium, potassium (less affected than sodium and chloride), chloride (if triglyceride concentrations are high, e.g. > 600 mg/dL).
Unknown if increased or decreased: ALT, AST, bile acids.
≥500 Falsely increased:  Direct bilirubin, TIBC.
Falsely decreased
: sodium, potassium (less affected than sodium and chloride), chloride (if triglyceride concentrations are high, e.g. > 600 mg/dL).
Unknown if increased or decreased: Albumin, ALT, AST, bile acids.
≥1000 Falsely increased:  Direct bilirubin, TIBC.
Falsely decreased
: sodium, potassium (less affected than sodium and chloride), chloride (if triglyceride concentrations are high, e.g. > 600 mg/dL).
Unknown if increased or decreased: Albumin, ALT, AST, bile acids, calcium, cholinesterase, CK, glucose, phosphate, urea nitrogen. 
 ≥1500 Falsely increased:  Direct bilirubin, TIBC.
Falsely decreased:
Bicarbonate, sodium, potassium (less affected than sodium and chloride), chloride (if triglyceride concentrations are high, e.g. > 600 mg/dL).
Unknown if increased or decreased:
Albumin, ALT, AST, bile acids, calcium, cholinesterase, CK, fructosamine, glucose, phosphate, total bilirubin and urea nitrogen. 
Effect on other analytes: Unknown.

 

Hemolysis index

Hemolysis tubes

Different degrees of hemolyzed serum

The number reported under Hemolysis on our chemistry panels is a semi-quantitative measurement of the concentration of free hemoglobin in mg/dL. The image to the right shows a visual representation of how these values correspond to visible hemolysis in plasma samples and the table below correlates the index value to this gross appearance.

Guide to Hemolysis Index
Hemolysis index Gross appearance
<20 No hemolysis
20-100 Slightly hemolyzed (pink tinged)
100-300 Moderately hemolyzed (red)
>300 Markedly hemolyzed (dark red)

 

Free hemoglobin in the sample has little or no effect on most assays unless the level is very high (>300 hemolytic index). The guidelines below indicate how hemolysis may affect sample results from our current chemistry analyzer. These guidelines are derived from manufacturer’s recommendations and is based on data from spiking human samples with hemoglobin. These values may not apply to samples from animals and there are important species differences (for example, a high potassium is expected in hemolyzed samples from horses, camelids, pigs and sheep because potassium concentrations in the erythrocytes of these species are higher than in serum – this is not true for most breeds of dogs, other than Asian breeds, or several breeds of cattle). These guidelines should always be used in conjunction with clinical signs and knowledge of the underlying disease processes as guides for interpreting laboratory data and determining the accuracy of reported laboratory results. Species-dependent changes have been published, however these will always be method- and analyzer-dependent and are not broadly applicable to results. For more on species-dependent changes, refer to the interference section of test basics.

Effect of hemolysis on chemistry results (guidelines)
Hemolysis index Possible effect on results
≥19 Release of RBC constituents (false increase): LDH (in dogs, in our experience) 
≥40 Release of RBC constituents (false increase): LDH (in dogs, in our experience), TIBC.
≥100 Release of RBC constituents (false increase): LDH (in dogs, in our experience), TIBC, potassium (species-dependent – horses, camelids, cats and some ruminant breeds), CK (constituents in red blood cell membranes participate in the CK reaction)
Unknown if increased or decreased: Fructosamine.
≥200 Release of RBC constituents (false increase): AST, iron, LDH (in dogs, in our experience), TIBC, potassium (species-dependent – horses, camelids, cats and some ruminant breeds), CK (constituents in red blood cell membranes participate in the CK reaction)
False decrease: GGT.
Unknown if increased or decreased: Fructosamine.
≥300 Release of RBC constituents (false increase): AST, iron, LDH (in dogs, in our experience), TIBC, potassium (species-dependent – horses, camelids, cats and some ruminant breeds), CK (constituents in red blood cell membranes participate in the CK reaction).
False increase: Phosphate.
False decrease: GGT.
Unknown if increased or decreased: Fructosamine.
≥500 Release of RBC constituents (false increase): AST, iron, LDH (in dogs, in our experience), TIBC, potassium (species-dependent – horses, camelids, cats and some ruminant breeds), CK (constituents in red blood cell membranes participate in the CK reaction). 
False increase: Phosphate, total protein.
False decrease: Amylase, direct bilirubin, GGT, NEFA (by an average of 0.05 mEq/L, range of 0.03 to 0.07 mEq/L).
Unknown if increased or decreased: Fructosamine.
≥600 Release of RBC constituents (false increase): AST, iron, LDH (in dogs, in our experience), TIBC, potassium (species-dependent – horses, camelids, cats and some ruminant breeds), CK (constituents in red blood cell membranes participate in the CK reaction). 
False increase: Phosphate, total protein.
False decrease: Amylase, bicarbonate, direct bilirubin, GGT, NEFA (by an average of 0.05 mEq/L, range of 0.03 to 0.07 mEq/L).
Unknown if increased or decreased: Fructosamine.
≥700 Release of RBC constituents (false increase): AST, iron, LDH (in dogs, in our experience), TIBC, potassium (species-dependent – horses, camelids, cats and some ruminant breeds), CK (constituents in red blood cell membranes participate in the CK reaction).
False increase: Cholesterol, cholinesterase, phosphate, total protein, triglycerides.
False decrease: Amylase, bicarbonate, direct bilirubin, GGT, NEFA (by an average of 0.05 mEq/L, range of 0.03 to 0.07 mEq/L).
Unknown if increased or decreased: Fructosamine.
≥800  Release of RBC constituents (false increase): AST, iron, LDH (in dogs, in our experience), magnesium, TIBC, potassium (species-dependent – horses, camelids, cats and some ruminant breeds), CK (constituents in red blood cell membranes participate in the CK reaction).
False increase: Cholesterol, cholinesterase, phosphate, total protein, triglycerides.
False decrease: Amylase, bicarbonate, direct bilirubin, GGT, NEFA (by an average of 0.05 mEq/L, range of 0.03 to 0.07 mEq/L).
Unknown if increased or decreased: Creatinine, fructosamine, total bilirubin.
 ≥1000  Release of RBC constituents (false increase): AST, iron, LDH (in dogs, in our experience), magnesium, TIBC, potassium (species-dependent – horses, camelids, cats and some ruminant breeds), CK (constituents in red blood cell membranes participate in the CK reaction).
False increase: Cholesterol, cholinesterase, phosphate, total protein, triglycerides.
False decrease: Amylase, bicarbonate, direct bilirubin, GGT, NEFA (by an average of 0.05 mEq/L, range of 0.03 to 0.07 mEq/L).
Unknown if increased or decreased: Creatinine, fructosamine, total bilirubin.
Effect on other analytes: Unknown.

Icteric index

The number reported under icterus is an estimation of the total bilirubin concentration in mg/dL rounded to the nearest whole number. 

The icterus index can be used to determine if there is hyperbilirubinemia, i.e. if the total bilirubin is increased, the icteric index should closely match the value seen. For example, a serum sample from a dog that has an Icterus index result of “4” has a total bilirubin concentration of approximately 3.5 to 4.4 mg/dL, which indicates that the dog has hyperbilirubinemia. However, if there is a mismatch, e.g. the total bilirubin is 2 mg/dL and the icteric index is 0, the total bilirubin result is falsely increased. This type of artifact is seen in the following situations:

  • Underfilling of heparin anticoagulant tubes: If heparin (green top) anticoagulant tubes are <3/4 full with blood, a falsely increased total bilirubin (mostly indirect bilirubin) may be seen.
  • Species that normally have lightly colored serum (e.g. birds, ruminants): Carotenoids in the diet (and serum) will contribute to the icteric index (but not the total bilirubin value).

The guidelines below indicate on how tests are affected by icterus with our chemistry analyzer. These guidelines are derived from manufacturer’s recommendations and is based on data from spiking human samples with conjugated or unconjugated bilirubin. These values may not apply to samples from animals and there may be species differences. These guidelines should always be used in conjunction with clinical signs and knowledge of the underlying disease processes as guides for interpreting laboratory data and determining the accuracy of reported laboratory results.

Effect of icterus on chemistry results (guidelines)
Icteric index Possible effect on results
≥6 Unknown if increased or decreased: Fructosamine.
≥11 Falsely decreased:  Total protein (Garner et al 2014, Gupta & Stockham 2014), triglycerides.
Unknown if increased or decreased
: Fructosamine.
≥16 Falsely decreased:  Cholesterol, creatinine, total protein, triglycerides.
Unknown if increased or decreased: Fructosamine.
≥20 Falsely decreased:  Cholesterol, creatinine, GGT, total protein, triglycerides.
Unknown if increased or decreased: Fructosamine.
 ≥40 Falsely decreased:  Cholesterol, creatinine, GGT, total protein, triglycerides, uric acid.
Unknown if increased or decreased:
Fructosamine, phosphate.
  ≥50 Falsely decreased:  Cholesterol, creatinine, GGT, lipase, total protein, triglycerides, uric acid.
Unknown if increased or decreased:
Direct bilirubin, fructosamine, phosphate.
 ≥60 Falsely decreased:  Cholesterol, creatinine, GGT, total protein, triglycerides, uric acid.
Unknown if increased or decreased:
Direct bilirubin, fructosamine, phosphate.
Effect on other analytes: Unknown.

 

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