Skip to content
SearchProgress
Display

Display

Theme
Density
Text size

Sign in

Add your earlier progress to your account?

Study progress is waiting to be saved

Electrodes, blood gases, luminescence and light scatter

15 min

  • Explain why an indirect ion-selective electrode (ISE) can show pseudohyponatremia
  • Distinguish measured blood-gas values from calculated values
  • Distinguish excitation-driven fluorescence from chemical light production
  • Distinguish turbidimetric attenuation from nephelometric scatter

Read the full reference

Try first

Try first

A serum specimen with severe hyperlipidemia is tested for sodium by two methods. Which method can report a falsely low sodium?

The next section explains it.

The next section explains it.

Right. The next section explains why.

The next section explains it.

Get the idea

Direct and indirect electrodes

An ion-selective electrode (ISE) responds to ion activity in the water phase of the specimen.1,2

  • A direct ISE measures undiluted plasma or whole blood, so it reads sodium in the water phase alone.
  • An indirect ISE dilutes a fixed volume of plasma with buffer and calculates sodium per volume of whole plasma. The calculation assumes plasma is about 93% water.

Severe hyperlipidemia or hyperproteinemia lowers the water fraction. The fixed plasma volume then carries less water and less sodium into the diluent, and the indirect result is falsely low. This artifact is pseudohyponatremia. A direct ISE on the same specimen stays close to the physiologic value.2,3

Measured and calculated blood-gas values

A blood gas analyzer measures pH, pCO2 and pO2 with electrodes. Co-oximetry measures the hemoglobin fractions by absorbance at several wavelengths. Bicarbonate is calculated from pH and pCO2.2,4

A saturation calculated from pO2 and an assumed dissociation curve uses no hemoglobin measurement. It cannot show carboxyhemoglobin or methemoglobin. In carbon monoxide poisoning the pO2 and the calculated saturation can look normal as the measured oxyhemoglobin falls.2,4

Light from a lamp or from a reaction

In fluorescence, a fluorophore absorbs excitation light and emits light of a longer wavelength. The difference is the Stokes shift. The detector sits away from the excitation path, often at 90 degrees.1,2

In chemiluminescence, a chemical reaction makes the excited product, and a photomultiplier tube reads the light it gives off. The analyzer has no excitation lamp. Low or high signal comes from reagent or substrate deterioration, timing error, incomplete washing, carryover or detector saturation.2

Scatter read in line or off axis

Particles in suspension scatter light, and the detector's position decides what is measured.1,2

MethodDetector positionAs concentration rises
TurbidimetryIn line with the beamTransmitted light falls
NephelometryOff axis, commonly 30 to 90 degreesScattered light rises from a dark background

Nephelometry generally detects lower concentrations, because it measures a small signal against a dark background.2

References
  1. Bishop ML, Fody EP, Van Siclen C, Mistler JM, Moy M. Clinical Chemistry: Principles, Techniques, and Correlations. 9th ed. Jones & Bartlett Learning; 2023.
  2. Rifai N, Chiu RWK, Young I, Burnham CAD, Wittwer CT, eds. Tietz Textbook of Laboratory Medicine. 7th ed. Elsevier; 2023.
  3. Fortgens P, Pillay TS. Pseudohyponatremia revisited: a modern-day pitfall. Arch Pathol Lab Med. 2011;135(4):516-519. doi:10.5858/2010-0018-RS.1
  4. Clinical and Laboratory Standards Institute. Blood Gas and pH Analysis and Related Measurements. 2nd ed. CLSI guideline C46-A2. Clinical and Laboratory Standards Institute; 2009. Archived and retained as technically valid. Accessed September 26, 2026. https://clsi.org/shop/standards/c46/

Watch one

An arterial specimen from a patient brought in from a house fire gives these results on a blood gas analyzer with co-oximetry:

  • pH 7.36, pCO2 38 mm Hg and pO2 96 mm Hg
  • calculated oxygen saturation 97%
  • oxyhemoglobin fraction (FO2Hb) 71%, carboxyhemoglobin 26%, methemoglobin 0.6% and deoxyhemoglobin 2.4%

Which results describe the patient's oxygen-carrying hemoglobin?

  1. Sort the results: pH, pCO2 and pO2 come from electrodes, the four hemoglobin fractions come from co-oximetry, and the saturation is calculated.

    Measured and calculated values rest on different evidence, and the report shows both.

  2. Read the pO2: 96 mm Hg sits inside the arterial reference interval of 75 to 100 mm Hg.

    The partial pressure of oxygen measures dissolved oxygen and says nothing about which hemoglobin species carry it.

  3. Set the calculated saturation of 97% aside, because no hemoglobin was measured to produce it.

    The calculated saturation comes from the partial pressure of oxygen and an assumed curve, so it cannot see carboxyhemoglobin either.

  4. Read the measured fractions: carboxyhemoglobin is 26%, and oxyhemoglobin is only 71% of total hemoglobin. The four fractions add to 100%.

    Co-oximetry resolves every species the instrument supports, carboxyhemoglobin included.

  5. Report the measured fractions, and follow your laboratory's procedure for notifying a high carboxyhemoglobin.

    A high carboxyhemoglobin may need a prompt call under the laboratory's notification list.

The co-oximetry fractions describe the hemoglobin: oxyhemoglobin 71% and carboxyhemoglobin 26%. The calculated saturation of 97% cannot show carboxyhemoglobin.

Your turn

Problem 1 of 3

Why can severe hyperlipidemia cause a falsely low sodium result with an indirect ion-selective electrode (ISE)?

Incorrect. Membrane deposits cause drift or slow response on any electrode, direct or indirect. The indirect artifact comes from dilution. Lipid displaces plasma water, so the fixed plasma volume carries less sodium into the diluent.

Incorrect. An ion-selective electrode measures a potential difference at negligible current, so turbidity has no path into the reading. Lipid lowers the indirect result by displacing plasma water.

Correct. Indirect ISE dilutes a fixed plasma volume and assumes about 93% water. Excess lipid lowers that fraction, so less water and less sodium enter the diluent and the calculated sodium is falsely low. Direct ISE reads the undiluted water phase and stays close to the physiologic value.

Hint
  1. Picture what goes into the diluent when the analyzer takes its fixed volume of plasma.
  2. Lipid takes up space in the plasma that water would normally fill.
  3. The indirect calculation assumes a water fraction. Ask what happens when the real fraction is lower.

Review Electrochemical measurements

Problem 2 of 3

A nephelometer measures immunoglobulin G through antigen-antibody complexes. Within the measuring interval, what does its detector record as the IgG concentration rises?

The off-axis detector collects scattered light. More complexes scatter more light toward it, so the signal rises from a dark background.

Falling light from a blocked beam is what an in-line turbidimetric detector records. A nephelometer's detector sits off axis and receives more light as scatter increases.

Treated transmitted-light loss as off-axis scatter

Turbidimetry reads the light lost from the transmitted beam with an in-line detector. Nephelometry reads light scattered to an off-axis detector. As concentration rises, transmitted light falls and scattered light rises from a dark background, which is why nephelometry detects lower concentrations. Confusing the two reads the signal change in the wrong direction.

A shift to a longer wavelength is emission from a fluorophore. Scattered light keeps the wavelength of the source.

Hint
  1. Find where the nephelometer's detector sits relative to the beam.
  2. More complexes scatter more light away from the straight path.

Review Turbidimetry and nephelometry

Problem 3 of 3

After a new reagent pack is loaded on a chemiluminescent immunoassay analyzer, both control levels read well below their means. A colleague suggests replacing the excitation lamp. What do you check first?

A chemiluminescent assay makes its light by a chemical reaction. The analyzer has no excitation lamp to fail.

Looked for an excitation lamp in a chemiluminescent assay

Chemiluminescence produces its excited product by a chemical reaction, so the analyzer has no excitation source to fail. Troubleshooting a lamp or excitation filter misses the causes of low or high signal in these assays: reagent or substrate deterioration, timing error, incomplete washing, carryover, or detector saturation.

A Stokes shift belongs to fluorescence, where light is absorbed and then emitted. A chemiluminescent assay has no excitation light to separate from emission.

Low signal in a chemiluminescent assay comes from the chemistry that makes the light and the steps around it. The problem started with the new pack, so its reagents and substrate come first.

Path length matters for absorbance measured through a cuvette. A chemiluminescent assay counts photons emitted by the reaction.

Review Fluorescence and chemiluminescence

Use it

  • A 44-year-old woman in the emergency department (MRN 0061437) has serum electrolytes and a whole-blood blood gas panel drawn together at 09:40.
  • The serum is milky. Her triglycerides, measured after dilution, are 5,200 mg/dL.
  • The chemistry analyzer measures electrolytes by indirect ISE.
  • The blood gas analyzer measures whole-blood sodium by direct ISE.
  • Both analyzers' controls were acceptable this morning.
TestResultPreviousReference intervalFlag
Sodium (serum, indirect ISE)128 mmol/L138 mmol/LSep 23, 08:15135–145 mmol/LLow
Chloride (serum, indirect ISE)95 mmol/L103 mmol/LSep 23, 08:1598–107 mmol/LLow
Potassium (serum, indirect ISE)3.9 mmol/L4.1 mmol/LSep 23, 08:153.6–5.2 mmol/L
Sodium (whole blood, direct ISE)139 mmol/L135–145 mmol/L

Specimen: H 10, L 850, I 2. Serum and heparinized whole blood, both collected 09:40 from the same venipuncture.

Decision 1 of 2

Why do the two sodium results differ?

Deposits on an electrode cause drift or a slow response. They do not add sodium, and the blood gas analyzer's controls were acceptable.

The indirect method dilutes a fixed volume of plasma and assumes about 93% water. This milky serum holds less water, so the calculated sodium comes out low. The direct electrode reads the water phase and gives 139 mmol/L.

Both specimens came from the same venipuncture at 09:40. The difference lies in the methods.

The direct electrode measures undiluted whole blood and reads sodium in the water phase. Lipid does not lower that reading.

Expected direct ISE to show the indirect sodium artifact

Direct ISE reads sodium activity in the water phase of undiluted plasma, so excess lipid or protein does not lower it. When an indirect sodium is low and a direct sodium on the same specimen is within the reference interval, the low indirect result is pseudohyponatremia caused by the lipid or protein.

Review Electrochemical measurements

Decision 2 of 2

What do you do with the serum sodium and chloride?

The indirect method already dilutes the specimen. Extra dilution keeps the same low water fraction in the plasma that was sampled, so the result stays falsely low.

A sodium of 128 mmol/L in the chart reads as hyponatremia the patient does not have. It could be acted on.

The indirect sodium and chloride share the same dilution error. The direct ISE result reflects her plasma water, and the comment explains why the chemistry analyzer's electrolytes were not reported.

A repeat on the same indirect method repeats the same error and gives about the same low results.

Review Electrochemical measurements

The clue that settles this case is the pair of sodium results from one draw. The indirect sodium is 128 mmol/L, and the direct sodium is 139 mmol/L, about 8% apart. The lipemia index is 850 and the triglycerides are 5,200 mg/dL, so lipid takes up enough of the plasma to explain the gap. The direct measurement shows her sodium within the reference interval.

Keep

Sources checked