Measuring with the e-BQC device and obtaining wrong results

MOST COMMON e-BQC MISTAKES: RESULTS WITH LOW REPRODUCIBILITY, ACCURACY AND ERROR MESSAGES

The e-BQC is a very sensitive device that uses electrochemistry to measure antioxidant capacity. This means that there are some factors to take in account and good practices to minimize the variation when doing replicates, and the display of error messages. These e-BQC mistakes are related mainly to three main keystones:

e-BQC Strips

The measurement takes place on the strips, so it is very important to take good care of them. The strips have several parts as can be seen here.

THE MISTAKES

  • Touching them with bare hands
  • Reusing the same strip
  • Causing  scratches to the active site
  • Strips with dust

THE SOLUTIONS

  • Wear clean gloves to handle the strips and never, even with clean gloves, touch the active site, where the sample is disposed on.
  • Use new strips for each measurement and never use the same strip to measure.
  • Maintain the strips in the container until used to avoid damage or dirt.
  • Avoid rubbing the strips against themselves or any other material.
  • Do not use any strip that seems damaged.
  • When pipetting the samples, do not touch the surface of the strip with the tip.
Most important parts of the strip

SAMPLE:

Some samples will present more reproducibility than others because it depends on their composition which will determine their suitability to measure with the e-BQC device.

THE MISTAKES

  • Lipidic composition
  • Not taking in account the pH
  • Heterogeneous samples
  • Low conductivity
  • High viscosity
  • Showing matrix effect
  • High protein content
  • Interferant components present in the sample

THE SOLUTIONS

  • Remove the organic diluents or dilute your sample with an aqueous buffer at least to a 1:2 ratio.
  • To avoid the pH effect, some samples may need to standardize the pH. Do not compare samples with very different pH values.
  • Work with homogeneous samples, if not possible, try to separate interferent substances.  Use higher volumes to measure because it will take a more representative sample of components.
  • Samples with low conductivity will require dilution with a buffer. 
  • For the matrix effect case a calibrate is recommended. The lower the slope of the calibrate, the higher the matrix effect. In the graphic a calibrate for a milk sample is plotted.
  • Avoid letting the sample in the strip for a long time, instead measure immediately. For protein extracts, use a buffer to dilute and test different concentrations to optimize your results.
  • Check the presence of exogenous or added antioxidants.

Antioxidants are very sensitive substances that because of their special properties, are oxidized with ease. For this reason, another factor to take in account is the sample handling.

THE MISTAKES

  • Light exposure
  • Oxygen exposure
  • High temperature of storage
  • Long time of storage

THE SOLUTIONS

  • Keep your samples in opaque containers and avoid direct lighting.
  • Handle your samples with care. Do not mix by vortexing at high speeds, instead, use inversion and mixing with the pipette. Avoid bubbles when disposing the drop, one way to do that is not pulsing the second stop of the pipette.
  • Store samples at low temperatures (between -80ºC and -20ºC) to ensure prolonged stability. Before measuring, let the samples reach room temperature. Do not compare results of samples with different temperatures. Avoid multiple freeze-thaw cycles, instead, use aliquots.
  • As far as possible, use freshly obtained samples. If not possible, store at low temperatures (between -80ºC and -20ºC) and measure the sooner the better. Do not use samples that have been stored for a long time or inappropriately stored.
Sample with matrix interference

USAGE

After the sample and strips handling, a correct usage should be ensured for reproducibility and accuracy.

THE MISTAKES

  • Inadequate coverage
  • Too low sample volumes
  • Not changing tips between measurements
  • Using different sample volumes and comparing the results
  • Bubbles in the drop
  • Moving/vibrating working surface
  • Misleading statistics

THE SOLUTIONS

  • Follow the scheme as a guide for a good coverage, all the surface inside of the blue protector should contain sample. Ensure that the control zone is completely covered, preferably, start with this part to fill the active site of the strip.
  • Use more than 30 µL of sample, for example 50 µL. If the sample is very viscous, more volume should be used. If there is no sample volume limitation, 100 µL ensures complete strip coverage.
  • Always change tips between samples to avoid cross-contamination.
  • Be consistent with the volume chosen for all the tests that will be compared lately. For heterogeneous samples, use higher volumes.
  • Maintain the strips in the container until used to avoid damage. Avoid rubbing the strips against themselves or any other material. Do not use any strip that seems damaged. When pipetting the samples, do not touch the surface of the strip.
  • Avoid bubbles when disposing the drop, one way to do that is not pulsing the second stop of the pipette. This will cause the same amount of volume not to be dispensed but it will be the same error for every measurement.
  • Ensure that the surface in which you are working is smooth, horizontal and stable. Do not move the device or the table while measuring.
  • Use non-relative statistics.
Strip coverage guide

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