Research integrity (antibodies and chemicals)

Introduction

A large issue in reproducibility is the quality of our consumables that are often not at the standard required for research. Unfortunately, we always assume that our suppliers provide the right items that works as described in their brochures. This is unfortunately not true all the times. We have only one way to cope with this issue, validate also commercially sourced material, store and stock it properly, trace its origin appropriately.

We should also admit that we cannot dedicate all time and budget required for this activity. Therefore, always think about protocols and procedures that minimize waste of time and money while ensuring we use high grade materials.For materials such as salts and solvents, we of course will assume companies doing their basic duties. However, we need to pay attention to antibodies and drugs in particular.

Storage (draft)

Make sure you store all materials according to supplier guidelines. Occasionally, we have some material that does not work as it is supposed to. It is therefore important we keep track of lot number and also the particular lab stock, so that as soon as an issue is identified, stock and aliquotes can be withdrawn from usage. We are aware that tubes are sometimes small, but you can store individual aliquotes within a larger Falcon tube or in a sealable bag, where all information can be written. Keep track also in your lab book of this information. We are aware this is a tedious task and often superfluos, but – however, it is the only way to control issues of reproducibility and to purge not just bad materials, but also experiments that were executed with these.

[Open task] We have to establish a flexible, efficient, non time consuming inentory system that ensure tracability of materials and experiments.

Validation (draft)

Validation of materials is necessary. All materials generated in the lab will have to be validated within the lab. What we outsource to commercial suppliers is expected to be validated at source. However, do double-check how the supplier validated the material. In most cases, we can ensure that the experimental design is capable to capture the appropriate function of materials. For instance, if we use commercial, well-known MEK inhibitor and do a Western blot, we always have a treated and untreated sample and always blot for phosphorylated ERK to verify the effect of the treatment. When we do a knock-down, we always verify we actually knocked-down the target. Never run an experiment where you knocked down a protein and you did not verify the knock-down, for instance by Western blotting. There is a number of experiments where we do not or cannot do the same. For instance, long-term live cell imaging with microfluidics where we cannot run a parallel experiment. However, we should design also these experiments with validation in mind. When we use a cytotoxic agent, we know how much cell death we should achieve in a control. If we do not, the tratement did not work. However, we now reccomand to also run a test experiment in parallel, even if it is in a different format, to verify the effectivness of the treatment. For instance, when using a DNA damaging agent, you can use the same aliquotes and cells in a different chamber but similar condition, then staining for gH2AX. This type of controls will help you in troubleshooting, but also it will allow us to decide in a rigorous way if one experiment should be discarded because of issues with materials or execution, or if it might represent biological variability.

Antibodies

Western-blotting

Experimental conditions