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Application Information
Apply for laboratory accreditation.
Program Information
More information about the LabCert program.
Resources & Training
Tools to assist you in testing.
Contact information
For information on the Lab Certification program, contact:
Tom Trainor
Program chemist
Science Services / Laboratory Certification

DNR LabCert mailbox
Ultimate BOD header graphic

BOD equipment & supplies

BOD Incubator

BOD incubator
Thermostatically controlled at 20± 1°C. All light must be excluded to prevent photosynthesis during the 5-day incubation period.


Digital Models

dial type barometer
The laboratory temperature and barometric pressure must be recorded daily, particularly if the dissolved oxygen (DO) meter is calibrated using the water-saturated air or air-saturated water technique. NIST traceable electronic models are available that will monitor both temperature and barometric pressure are available.

Dial type models

dial type barometer
Dial type NIST traceable barometers are satisfactory for monitoring barometric pressure. All barometers (electronic and dial type) should be checked to verify they are functioning properly. The internet or local airport can provide information that maybe used to check laboratory barometers. See the calibration section of this document for information on how to check barometers.

BOD Bottles

Standard glass bottles

standard glass BOD bottles
Standard glass, 300 mL BOD bottles. Glass BOD bottles are most commonly used by laboratory since they may be washed and reused. Wash BOD bottles using technique described elsewhere in this document.

Disposable bottles

standard glass BOD bottles
Single use, plastic disposable 300 mL BOD bottles. These bottles are acceptable for use providing they are only used once and all quality control criteria are met.

Proper cleaning of BOD bottles

Use a good lab-grade non-phosphate detergent and bleach. Rinse thoroughly with tap water followed by distilled water. Allow to dry before storing. Always cover glassware and store in a clean, dry place.

Bleach-free alternate cleaning method

Danger! Chlorine gas.

DO NOT MIX hydrochloric acid (HCl) and bleach: It will produce poisonous chlorine gas!!!!.

Some folks prefer not to use bleach at all in cleaning their BOD labware, as even traces of bleach can kill "bugs" critical to the BOD test. Use a good laboratory grade non-phosphate detergent. Rinse thoroughly with tap water followed dilute hydrochloric acid (HCl) [10% solution; 100 mL hydrochloric acid (HCl) per liter of water]. Rinse again w/ tap water followed by distilled water. Allow to dry before storing. Always cover glassware and store in a clean, dry place.

Dilution water containers

glass carboy for storage of lab reagent water or dilution water
  • Store dilution water in all glass containers only. Any hard plastic will break down over time and leach BOD.
  • If dilution water is siphoned into BOD bottles from the container, only Teflon® or glass should be immersed in the water.
  • Use only C-flex® and latex rubber tubing to dispense the dilution water into BOD bottles.

glass carboy for storage of lab reagent water or dilution water
  • Bottom draw, aspirator bottles can also work well. Polypropylene bottles do, however, present a risk. As bottles are cleaned with bleach or acid, the plastic breaks down, posing a risk for leaching of BOD material. Glass bottles with glass aspirators are always preferable.

wrap glass carboys with reinforced tape for safety
  • For greatest safety, wrap large glass carboys with reinforced tape. This will minimize generation of dangerous glass shards if the bottle breaks.

Saturating BOD dilution water with oxygen

Use of airstones to aerate dilution water is NOT recommended.

As a general rule, NEVER put "fish tank" tubing or air stones directly in contact with dilution water. These items can leach oxygen demanding materials. Air stones can harbor bacteria which--unaccounted for--can cause excess DO depletion and subsequent high bias

in-line filter used to help purify air used to aerate dilution water
Saturating dilution water with oxygen can be source of contamination in BOD testing. Water can be aerated by lightly covering the opening of the glass storage container with link free tissue, cotton or foam rubber. Avoid using material that will slough-off into the dilution water. If a laboratory uses compressed air to aerate the dilution water, use an in-line filter (as shown) to prevent dust and oil from getting into the water. In-line filters are available from most scientific specialty companies.

Tubing types

Acceptable tubing types for use in BOD testing

photo: George Bowman, Wisconsin State Laboratory of Hygiene

Pipet types

Acceptable pipet types for use in BOD testing

photo: George Bowman, Wisconsin State Laboratory of Hygiene

Certified thermometers

A factory (or NIST) certified thermometer must be placed in the incubator to verify the temperature is maintained within the 20± 1°C tolerance each day during the 5-day incubation period (including weekends). These may also be used to demonstrate that the temperature of sample refrigerators and autosamplers is maintained at temperatures not to exceed 6°C (and samples cannot be frozen). Total immersion type incubator thermometers or electronic thermometers are satisfactory. Always record the temperature in a logbook or bench record.

Smaller thermometers in a sealed bottle are perfect for incubators and refigerators.
Smaller NIST traceable thermometers work well for documenting incubator temperatures. Rather than re-calibrating, these are usually replaced annually.
Partial immersion thermometers have an immersion line inscribed on them.
A partial immersion thermometer is needed to confirm the samples are in the 20± 3°C range prior to preparing the BOD dilutions. Be wary of the immersion line scribed on these thermometers.
Minimum-maximum thermometer
NIST traceable minimum/maximum or electronic recording thermometer are adequate for checking the temperature on weekends.

DO meter and DO probe

A dissolved oxygen (DO)meter and probe are critical items needed for the BOD testing.

Dissolved oxygen (DO) meter
Most new newer DO meters have a built-in barometers, thermometer and on-board software to simplify the calibration process.
Dissolved oxygen (DO)probes
While polarographic type DO probes used to be the most common style in use, they are quickly being out-paced by the newer, more rugged electro-optical (LDO, RDO) probes.

pH meter and pH paper

pH meter
A pH meter is the preferred way to verifying the sample pH is in 6.5-8.0 range prior to testing for BOD.
short range pH paper
Short range pH paper is an acceptable option as well. pH paper may not be used to measure pH for permit purposes, however.

Other supplies for BOD testing

  • Nutrient buffer solutions — commercially available
  • Sulfuric acid [H2SO4] solution (1N) for adjusting sample pH — available commercially
  • Sodium hydroxide [NaOH] solution (1N) for adjusting sample pH — available commercially
  • Glucose-glutamic acid (GGA) solution — available commercially
  • Nitrification inhibitor (if carbonaceous BOD is desired or allowed in the facility's NPDES permit) — available commercially (Hach formula 2533 © or equivalent)
  • Standard laboratory glassware
  • Bleach for sanitizing BOD bottles, dilution water storage containers and tubing used to dispense dilution water.
  • Hydrochloric acid [HCl] solution (10% of concentration) for adjusting pH or removing scale from BOD bottles.

Copyright 2006. University of Wisconsin Board of Regents.
Unauthorized use prohibited without the expressed written consent of the UW, State Laboratory of Hygiene and the DNR Laboratory Certification & Registration Program.

Last revised: Wednesday August 22 2012