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As a cleaning agent, sodium hydroxide saponifies fats and dissolves proteins. In general, it can solubilize precipitated proteins. Its hydrolysing power is enhanced by the presence of chlorine. The ability of sodium hydroxide to remove proteins and nucleic acids from chromatography media depends on the nature of the media, the sample, and sample contaminants that may interfere with the cleaning efficiency.

You can read the attached article for greater insight on functioning of NaOH as a sanitizer or cleaning agent.

Unless a chromatography resin cannot withstand it, like protein A resins, the standard concentration for NaOH treatment in biotech industryt is 0.5M for >= 1h. But I have seen protocolswith 1M NaOH, as well.

Yes, that's true. The ultrafiltration membranes in Minitan Milipore system are usually cleaned after protein concentrate with 1 M NaOH.

Hello Andreas, rereading your question, I am not sure you are actually covering a need in industry.

The cleanability for a GMP manufacturing process of a new biologic is tested and verified in a, so called, cleaning-validation. If the validation shows that something is not removed by the standard treatments, then more harsher methods are tried.

NaOH is cheap, one of the cheapest reagents used in biotech industry. Therefore, to make sure everything is clean and safe, the tendency will always be to go overkill. A few decades ago 0.1M NaOH was routinely used; now it is 0.5M or even 1M, and these high concentrations are not really needed for most steps in a manufacturing process. But its cheap and better safe than sorry. To have to redo a batch under GMP because of impurities costs easily $1M, often more.

However, of interest in the industry is how to deal with protein cakes on stainless steal walls. After cleaning a stainless steel tank that has seen protein, there are sometimes protein precipitates on the wall and they do not always dissolve and disappear with 0.5M NaOH treatment, even using hot NaOH. In some cases, they actually become hard plastic-like crusts and are become even more difficult to remove.

If you were to look into this problem and could find an easy solution ...

@ Achim Recktenwald

Hello Achim,

the need for industry (especially contract manufacturers) is to produce more batches in the same time. Long time frames for cleaning during product change over to assure absolute overkill costs not much money but if you are able to produce with the same equipment 25 instead of 20 batches whithin a year you will earn millions of euro. For overkill calculation you should know when a cleaning procedure reaches the target (e.g. acceptable carry over of TOC). If you show during cleaning validation that 3 hours of incubation with 0.5 M NaOH is valid and you do not know whether the result is reached after 15 minutes or 2 hours 55 minutes you cannot state that you have an overkill cleaning. Supposed 15 minutes are sufficient you would have an overkill cleaning with 30 minutes of incubation and win 2 hours and 25 minutes. May be you can use 1M NaOH, than you have an overkill cleaning with 15 minutes incubation too.

As contract manufacturer you cannot make studies for degradation with every monoclonal antibody or protein you produce for your clients. The API is expensive and studies are expensive too. For me it is important, if the degradation kinetics my be bracketed for proteins (e.g. number of peptide bonds in the molecule) and standard incubation times can be fixed as a minimum for different NaOH concentrations. Based on this times you develope your cleaning procedure which may be validated without knowing if it is overkill or not, but you avoid exorbitant cleaning time. Protein cakes are not the problem. Visual clean is always one acceptance criterion.

Hello Andreas: "As contract manufacturer you cannot make studies for degradation with every monoclonal antibody or protein you produce for your clients."

I would have to verify with our Regulatory group, but to my knowledge, cleaning validation, i.e., the ability to demonstrate the complete removal (down to very, very low limits) of a product that came in contact with surfaces of a piece of equipment, is a legal requirement for every molecule manufactured under GMP. Concluding that one mAb behaves like the next, is not valid; besides, I know of at least one instance, where this does not fit.

My assumption is that it is only partially a feature of the molecule, but dependent on the conditions (buffer, pH, conductivity, temperature, ...) during the exposure of the molecule to the surface.

Cleaning and cross-contamination are (almost) always a concern and topic during health authority inspections. As I said, using ~1h with 0.5M is quasi a standard in industry. If you suddenly change this to less, you will be scrutinized. And unless you can show this to be sufficient with undisputable data, you will be in trouble. For it to become a new standard, without testing from molecule to molecule, as you intend, you would have to show it to be sufficient with a large number of products and processes; and in the end you would still have to perform a cleaning validation.