Liofilchem® ONPG Test Kit, 30 Tests

Liofilchem® ONPG Test is a rapid in vitro diagnostic assay designed for the qualitative detection of β-galactosidase activity in bacterial isolates. The test is supplied as ready-to-use tubes containing a desiccated medium with ortho-nitrophenyl-β-D-galactopyranoside (ONPG), a chromogenic substrate used for microbial differentiation and identification, particularly among Gram-negative bacteria.

The ONPG Test is intended as an aid in bacterial identification workflows and must be used in conjunction with additional biochemical or diagnostic tests to achieve a complete and accurate identification.

Intended Purpose

The ONPG Test is intended for the detection of bacterial β-galactosidase activity from pure cultures. It supports the differentiation of lactose-fermenting, late lactose-fermenting, and non-lactose-fermenting organisms within routine diagnostic microbiology.

Principle of the Method

Lactose fermentation in bacteria depends on two enzymes:

• Permease, which transports lactose into the cell
• β-Galactosidase, which hydrolyzes lactose into glucose and galactose

Some Enterobacteriaceae (e.g. Escherichia coli, Klebsiella spp., Enterobacter spp.) produce both enzymes and are true lactose fermenters.

Late lactose fermenters (e.g. Citrobacter spp., Arizona spp.) produce β-galactosidase but lack permease.

True lactose non-fermenters (e.g. Salmonella, Shigella, Proteus, Providencia, Morganella) lack both enzymes.

ONPG enters the bacterial cell independently of permease. If β-galactosidase is present, ONPG is hydrolyzed to o-nitrophenol, producing a yellow color. Absence of color indicates no β-galactosidase activity.

Kit Contents

Each kit contains:

• 30 ONPG Test tubes (desiccated medium)

Order Reference: 88005

Pack Size: 30 tests

Reagents

Each tube contains a desiccated medium incorporating ortho-nitrophenyl-β-D-galactopyranoside (ONPG), a specific substrate for β-galactosidase detection.

Test Procedure Summary

1. Allow test tubes to reach room temperature.
2. Add 0.2 mL sterile physiological solution (0.85% saline) to the test tube.
3. Emulsify one or more well-isolated colonies from an 18–24 h culture.
4. Prepare a negative control tube containing saline only.
5. Incubate at 35 ± 2°C for up to 24 hours.
6. Examine for color change after 4 hours and up to 24 hours.

Positive and negative controls should be tested concurrently.

Interpretation of Results

• Positive reaction: Yellow color development
• Negative reaction: No color change

Quality Control

Performance testing using these strains met established acceptance criteria.

Storage and Shelf Life

• Store at 2–8°C, protected from light
• Shelf life: 1 year
• Do not use beyond expiry date or if deterioration is observed

Applications

• Differentiation of lactose-fermenting and non-lactose-fermenting bacteria
• Identification of Enterobacteriaceae
• Detection of β-galactosidase activity
• Clinical and diagnostic microbiology laboratories

Limitations

• Not suitable for organisms producing intrinsic yellow pigmentation
• Inoculation should be performed from lactose-containing media
• Reaction time may vary depending on inoculum size

Safety and Regulatory Information

• For in vitro diagnostic (IVD) use
• For professional laboratory use only
• Operators must be trained
• Follow standard biosafety and waste disposal procedures
• Compliant with IVDR (EU) 2017/746

Liofilchem ONPG Test offers a reliable and straightforward method for detecting β-galactosidase activity in bacterial isolates. Supplied as ready-to-use test tubes, it integrates easily into routine microbiology workflows and supports accurate differentiation of lactose-fermenting, late lactose-fermenting, and non-fermenting Gram-negative bacteria.