Lateral flow assay troubleshooting and understanding molecular diagnostic membrane switch-out

It’s a common scenario... your new diagnostic assay prototype is almost working the way you want it when an issue arises with a key component, the diagnostic membrane. You’re going to have to switch it out with an alternative, perhaps from another supplier, and that’s going to create substantial delays. Or is it?

Switching to a different lateral flow assay membrane is never easy, but there are things you and your new supplier can do to make the switch as smooth as possible. Read on to find out more and discover key troubleshooting tips.

Interested in testing a nitrocellulose membrane for your lateral flow assay, request a free sample now!

Lateral Flow assay example with nitrocellulose membrane

Why might I need to switch-out a nitrocellulose membrane for a lateral flow assay?

Lateral flow membranes form an integral part of diagnostic assays. Changing the nitrocellulose membrane in the later stages of assay development requires a complete revalidation of the test system, so developers will only switch membrane if there is a good reason. Ideally, they would also select a membrane and supplier that will help minimize any delays to market.

Quality and supply chain issues are two of the most common reasons for changing a nitrocellulose membrane. You might find that the pore size on a production batch is inconsistent with that of the test batch, jeopardizing the performance of your entire lateral flow development assay. Or your supplier might run into manufacturing or logistical difficulties, prompting a membrane switch-out.

How can a supplier simplify membrane switch-out for lateral flow assays?

Switching to a different diagnostic assay membrane doesn’t need to be difficult, though it does require time and resources. Key decisions in any diagnostic membrane switch-out process include identifying a new supplier and selecting a material and grade that’s fit for purpose.

Every supplier will be able to show you their ISO certifications, but can they help you get your diagnostic assay to market as quickly as possible? The aim is to find a supplier that can demonstrate a history of delivering high-flow nitrocellulose membranes while matching the specifications you give them, supporting you from the first day to the last.

One thing to look out for, regardless of whether a nitrocellulose membrane switch-out is due to quality or supply issues, is whether this potential new supplier can provide accurate comparison tables and knowledgeable people to help in finding a suitable replacement assay membrane. They should know their products best, and be able to help you streamline the switch-out process.

If quality improvements are the primary driver, the million-dollar question is whether you have successfully identified the material property needing improvement. For example, when liquid flow is inconsistent or slow, does the solution lie in the material’s chemistry (e.g. hydrophilicity) or in its physical properties (e.g. density, pore size, etc.).

The supplier’s in-house experts can work with you to confirm the existing issue, find any potential challenges, and provide appropriate test samples. Working closely with an experienced supplier can relieve some of the burden on the lateral flow assay developer, simplifying the nitrocellulose membrane switch-out process.

What can I do to ease the process of nitrocellulose membrane switching?

  • Membrane material specifications: Raw material sources and proprietary additives for glass fiber and paper pads vary between suppliers, leading to differences in performance that might need some adjustments to equalize.
  • Assay antibody chemistry: Although the risk is small, antibody chemistry might need adjusting to make sure the later flow assay’s sensitivity and reliability is unaffected by differences in nitrocellulose membrane surfactant.
  • Other assay redevelopment: Exchanging the lateral flow test membrane will most likely require a redevelopment of the dispensing blocking buffer systems and the conjugate pad chemistry.

While you might think addressing these points could add further delays, each is manageable with a well-informed supplier, and well worth considering from the start.


What are some other lateral flow assay troubleshooting solutions?

Should you still run into issues during membrane switch-out or revalidation, here are a few common challenges and (potential) quick fixes our in-house experts have encountered and solved. Hopefully you find them useful.

Challenge Possible solution
>Uneven lines or dots Use membranes with different pore size
Reduce dispensing volume of reagent
Increase protein concentration of reagent
Check dispensing buffer composition
Check dispensing process
False positive signals Modify buffer in conjugate pad/solution (e.g. pH, salt concentration, or surfactant concentration)
Change conjugated protein
False negative signals Modify buffer in conjugate pad/solution (e.g. pH, salt concentration, or surfactant concentration)
Change conjugated protein
Use membrane with smaller pore size
Increase sample volume

Uneven liquid fronts of migrating sample

Check membrane shelf life
Use membrane with different/more surfactant
Check whether the relative humidity is very low
Increase surfactant concentration in conjugate pad

Looking to try a new membrane in your existing solution? Building a new lateral flow device? Get a free sample of our Fast Flow High Performance nitrocellulose backed membrane.

For Samples or to contact a Specialist