Why do we use BABB all the time?

About BABB

BABB is a tissue clearing method invented by Andrew Murray & Marc Kirschner in the 1980s. It involves putting dehydrated tissues in a 1:2 v/v mixture of benzyl alcohol/benzyl benzoate, hence the abbreviation.

Why do we choose BABB in our standard pipeline?

BABB is the most efficacious and fastest tissue clearing agent and can clear the toughest tissues. It can clear a whole adult mouse, and any attempts to add things to it simply decrease its clearing power (Cai et al. Nat Neurosci. 22, 317–327 (2019). doi.org/10.1038/s41593-018-0301-3). 

It is also non-destructive to tissues. Tissues after BABB clearing can go for RNA transcriptomics (Yau et al. Nat Commun. 15, 10888 (2024). doi.org/10.1038/s41467-024-55248-0) and even proteomics (Bhatia et al. Cell. 185, 5040-5058.e19 (2021). doi.org/10.1016/j.cell.2022.11.021). 

Hence, for clinical use, BABB gives the best hope with stable performance, robust clearing, and molecular-level preservation. 

Is tissue shrinkage an issue?

The tissue shrinkage results from tissue dehydration during the alcohol dehydration steps. Throughout the history of histology, tissues are dehydrated before being embedded in paraffin wax for sectioning, undergoing repeated dehydration-rehydration cycles. Therefore, there is little problem with referencing the literature morphological data.

BABB is not toxic 

Contrary to the common paranoia, both benzyl alcohol and benzyl benzoate are not toxic. Both benzyl alcohol and benzyl benzoate are medically approved on humans as skin medication, in contrast, ethyl cinnamate is not. 

If you search the web for the Safety Data Sheets, all 3 compounds have dermal toxicity of LD50 in the range of 4000mg/kg in rats. 

BABB is not corrosive

Corrosive means it damages skin and tissues on contact. Per its use as dermal medication and tissue clearing agent, BABB is not corrosive. BABB also doesn’t corrode metals. The widespread misconception probably arose because BABB dissolves many plastics and paints, including polycarbonate and polystyrene, commonly used for Petri dishes, confocal dishes plastic parts, glues and resins that coat objectives, etc. Since humans are only full of microplastic but not fully plastic, BABB is not corrosive to humans (but can be for clothes, though a lot of things do, to be fair).

Polypropylene and polyethylene (LDPE or HDPE), that make up pipette tips, microcentrifuge tubes, and conical tubes are all safe indefinitely.

The above description is also true for ethyl cinnamate.

What’s with the smell of BABB?

Aerial oxidation sometimes produces benzaldehyde from benzyl alcohol, which has an almond smell. It is a common food flavouring and is literally produced by almonds during metabolism. Ethyl cinnamate also has a distinct flavour and is used as a fragrance and food flavouring as well.