Sterilization indicators: chemical, biological and Bowie-Dick tests

Sterilization indicators are the tests that show whether a sterilization cycle reached the conditions needed to kill microorganisms. There are three kinds: physical monitors on the sterilizer, chemical indicators that change color, and biological indicators containing bacterial spores. A Bowie-Dick type test is a special chemical test of air removal in prevacuum steam sterilizers. Facilities use all of them together.

The three types of sterilization monitoring

No single test proves a pack is sterile. Sterilization assurance comes from layers of evidence, each answering a different question.

Release decisions combine these. A load is released when physical readings are correct and the chemical indicators have passed; loads containing implants are held until the biological indicator result is known, except in documented emergencies.

Chemical indicators for sterilization: the six ISO 11140 types

ISO 11140-1 classifies chemical indicators by what they respond to. Knowing the types helps you buy the right sterilization indicator strips for each purpose.

• Type 1, process indicators. Autoclave tape, printed indicators on pouches and labels. They only distinguish processed from unprocessed items and go on the outside of every pack.
• Type 2, indicators for specific tests. The Bowie-Dick type test and similar air-removal tests.
• Type 3, single-variable indicators. Respond to one variable, such as a temperature reached. Rarely used in modern practice.
• Type 4, multi-variable indicators. Respond to two or more critical variables, such as time and temperature.
• Type 5, integrating indicators. Respond to all critical variables and are designed to perform in correlation with a biological indicator over a range of cycles.
• Type 6, emulating indicators. Matched to one specific cycle, for example 134°C for 3.5 minutes or 121°C for 15 minutes, and only pass when the full parameters of that cycle are reached.

Place internal chemical indicator strips in the area of each pack hardest for steam to reach, typically the center of a tray or between layers, and check the result when the pack is opened at the point of use. Store strips as the manufacturer instructs, because heat and humidity can affect them before use.

Chemical indicators examples by method exist for steam, dry heat, ethylene oxide and hydrogen peroxide. Use only indicators labeled for the process and cycle you run; a steam indicator tells you nothing about a hydrogen peroxide cycle.

Bowie-Dick and air-removal tests

Air trapped in a pack is the most common cause of steam sterilization failure in prevacuum machines. The Bowie-Dick type test detects it.

How it is used:

1. Run a warm-up cycle, or test when the sterilizer is at operating temperature.
2. Place the test pack alone, horizontally, in the empty chamber, usually on the bottom shelf above the drain.
3. Run the Bowie-Dick cycle, typically 134°C for 3.5 minutes.
4. Remove the indicator sheet and check for a uniform color change across the whole sheet.

A paler or unchanged center shows that air or non-condensable gases remained in the pack. Common causes are a chamber or door gasket leak, a weak vacuum pump, wet or poor-quality steam, or excess non-condensable gases in the steam supply. Take the sterilizer out of use and investigate before processing patient loads.

The original test used a towel pack; ready-made test packs and electronic test systems are now common and are covered in the ISO 11140 series. Small sterilizers that process hollow instruments may also use a hollow load process challenge device, often called a helix test, to show steam reaches the inside of a narrow lumen. Gravity displacement sterilizers do not use a Bowie-Dick test, because they do not have active air removal.

Biological indicators for steam sterilization

Biological indicators (BIs) are the only test that directly measures microbial kill. Each one carries a known population of highly resistant bacterial spores, commonly around 10^5 to 10^6, chosen because they are harder to kill than the microorganisms normally found on instruments. Requirements are set out in the ISO 11138 series.

Most steam biological indicators used in hospitals and clinics are self-contained vials: a spore carrier and a sealed ampoule of growth medium in one unit. After the cycle the ampoule is crushed and the vial is incubated, typically at 55–60°C for G. stearothermophilus. A color change or fluorescence signal shows surviving spores. Conventional systems read in 24 to 48 hours; rapid-readout systems detect an enzyme linked to spore survival and give results in minutes to a few hours.

Every batch of BIs tested should be accompanied by an unprocessed control from the same lot, incubated the same way, to prove the spores were viable. A test with a negative control is invalid.

Process challenge devices and test placement

A biological or chemical indicator is usually placed in a process challenge device (PCD), a test pack or tray built to be at least as hard to sterilize as the most difficult load the sterilizer processes. Hospitals often use commercial test packs or build a representative tray following their national guidance.

Place the PCD in the coldest or least accessible spot of the chamber, which for most steam sterilizers is on the bottom shelf near the door and drain, unless the manufacturer specifies otherwise. Run it with a full load for routine testing; some protocols call for an empty chamber after repairs.

How often should sterilization indicators be used?

Frequencies vary by country, but a common baseline, consistent with ANSI/AAMI ST79 in US hospitals, is:

• Every cycle: review physical printouts or data logs and sign off.
• Every pack: external process indicator, plus an internal chemical indicator.
• Every day the sterilizer is used: Bowie-Dick type test for prevacuum sterilizers, before the first load.
• At least weekly, preferably daily: biological indicator in a PCD.
• Every load with an implant: biological indicator, with the load quarantined until the result is known where possible.
• After installation, relocation, major repair or a failure: qualification testing, commonly three consecutive cycles with negative biological indicators, plus three Bowie-Dick tests for prevacuum machines.

Dental offices commonly spore test each sterilizer at least weekly. Follow the regulations, accreditation requirements and sterilizer IFU that apply to you.

Record-keeping and recalls

Indicator results are only useful if they are traceable. For each load, record the sterilizer, cycle number, date, operator, contents, physical readings and indicator results. Label every pack with the same load identification, so that a failed test can be traced to every pack processed since the last good result.

Write a recall procedure before you need it. It should say who decides on a recall, how items are retrieved, how patients are assessed when items have already been used, and how the sterilizer is returned to service.

Choosing sterilizers and supplies that support monitoring

When buying a steam sterilizer, check that it has test programs, a printer or data export, and stable cycles that indicators can verify. Keling's range spans benchtop units such as the KL-ST001 Portable Pressure Steam Sterilizer (18 L and 23 L, 134°C), vertical KL-ST002 Vertical Pressure Steam Sterilizer models from 35 to 100 L, and horizontal KL-ST004 Horizontal Pressure Steam Sterilizer chambers from 150 to 400 L. Ask for the cycle list, test programs and instructions for use of the exact model you plan to buy.

Perforated stainless steel trays such as the KL-TC076 Stainless Steel Sterilization Tray help steam reach instruments in a wrapped set; indicator placement should follow the same tray layout every time.

For more on choosing the machine, read our autoclave and steam sterilizer guide, and for the wider context of methods, see sterilization vs disinfection. Monitoring is one stage of the full sterile processing workflow.

This guide is educational and intended for trained staff. Always follow local protocols, your facility's policies and the instructions for use of each sterilizer and indicator.

Next steps

Set a written monitoring schedule for each sterilizer, buy indicators matched to your exact cycles, and keep traceable load records. To equip a clinic, laboratory or sterile processing department, browse Keling's laboratory and sterilization equipment and send your requirements through the quote form.