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> There was nothing to stop the smell of urine, and other volatiles, such as outgassing from plastics, from accumulating within the chamber.

The paper states that the substrate and changing schedule had been shown to keep ammonia to low levels.

> There's no reason to think the difference in noise was negligible...

Actually, there is. A good 80 mm fan can push more than 10 cubic feet per minute at less than 10 dBA. That's far quieter than a human whisper or rustling leaves. It's comparable to the sound of a pin dropping. If mice are adversely affected by that, I don't see how they could survive the wild.

> Plus, there may well have been other effects of the chamber, such as different light levels, which could also have unknown health effects.

That seems like pure speculation.

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What about things other than ammonia?

And did they have a "good" fan, or a bad fan?

The point of making the environment for the exposure and control groups be as near identical as possible is to equalize not only the factors you thought of, but also the factors you didn't think of.

It's the idea that there were no important differences between exposure and control groups, despite their obviously different living environments, that is "pure speculation".

> What about things other than ammonia?

Certainly possible, but it seems more likely that the control group would have been exposed to something harmful from the lab environment air versus the CO2 group that was getting medical air.

> And did they have a "good" fan, or a bad fan?

The cheapest 80mm fan I can find on newegg.com is a $3.49 Zalman[1] that is 20-23 dBA. That's comparable to the sound of rustling leaves and far less than the sound of a quiet whisper (30 dBA) or a babbling brook (40 dBA). Someone would have to go out of their way to find an 80mm fan that is so bad that it would be louder than a natural environment.

> The point of making the environment for the exposure and control groups be as near identical as possible is to equalize not only the factors you thought of, but also the factors you didn't think of.

I completely agree. I don't recall seeing a paper I couldn't poke holes in though. The fact that their methods were imperfect does not make the study worthless. They found a modest effect for the given methods. It suggests there may be merit to follow-on studies, which would likely use different methods. That's a useful result that has added new information.

[1] https://www.newegg.com/zalman-f1-led-case-fan/p/N82E16835118...

It’s too late to edit my previous comments, but I should mention that I actually agree with your statements about not being able to draw valid conclusions and that they should have used the same kind of chamber; however, I believe the methods were reasonable enough to provide some value.

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