Interferences in the LAL test: solutions

The LAL test allows you to detect the presence of endotoxin in pharmaceutical products, dialysis water, medical devices, etc. but to guarantee its success it is essential to avoid contamination and interference, which is why consumables free of endotoxin (pyrogenic) and certified non-interfering are used.

The most common sources of interference are:

• Inadequate LAL test consumables

• Water for LAL test not adequate

• Characteristics of the product to be analysed

LAL test consumables

Glass tubes

They are very important as they are used in all phases of the test: sampling, dilutions and finally the actual test.

The sampling and dilution tubes are made of borosilicate. The test tubes are made of borosilicate or soda glass and for the choice it is necessary to follow the instructions of the manufacturer of the LAL reagent. Borosilicate is a neutral glass which, unlike soda glass, does not release ions. The tubes can be used several times, after having depyrogenated them with dry heat.

Glass pipettes

Pyrogenic pipettes are usually made of borosilicate, certified with an endotoxin content of less than 0.001 EU/ml.

Plastic consumables

We are talking about containers, test tubes, pipettes, tips and microplates. It is necessary to use certified accessories or verify that the plastic is endotoxin-free and non-interfering in order to use it in the LAL test.

There are certified pyrogenic plastic products on the market with an endotoxin content of less than 0.001 EU/ml, but more often with a content of less than 0.005 EU/ml or 0.5 EU/device.

Water for LAL test

This is a fundamental element for the success of the test. The water for the LAL test must also be pyrogen-free and free of interfering factors.

The methods used for the depyrogenation of water are distillation (at least triple) and reverse osmosis. LAL water is usually certified with an endotoxin content of less than 0.001 EU/ml and can be bottled in glass or plastic.

Possible interfering substances present in water are:

Inhibiting substances

I.1) Ions such as chlorine, iron, aluminum and copper: present in mains water.
I.2) Substances that affect pH: pH problems are common after cleaning the system and after regeneration of the resins.

I.3) Organic substances: formalin or degraded resin.

Enhancing substances

S.1) LAL Reactive Material: glucans derived from cellulose filters

Product features

There are many factors that influence the LAL test and which can be overcome by applying certain strategies or using specific reagents. For many products, for example pharmaceutical or biotechnological, characterization is the phase in which the best strategy for performing the LAL test is defined, resolving any interferences, and which leads to the drafting of the SOP for routine analyses.

Factors with inhibitory effect

I.1) pH out of range 6.0 - 8.0
I.2) Presence of chelating agents
I.3) Excess of ions
I.4) Presence of proteins

*Factors with enhancing effect***

S.1) Presence of components of polysaccharide origin
S.2) Presence of protein aggregates

LAL test on Medical Devices

Plastic objects can show the same interferences seen in the previous sections. The LAL test is performed on the water used to rinse medical devices and usually the large volumes used dilute any interferences.