JISCMail - CCPEM Archives

My (non-expert) take on it is that, whether you use a 1/2 bit FSC criterion (asymptotic to 0.1716), advocated by Marin: https://www.biorxiv.org/content/biorxiv/early/2017/11/24/224402.full.pdf or a 0.143 FSC criterion justified by Peter, Tony and Richard in this appendix: https://www.sciencedirect.com/science/article/pii/S0022283603010222?via%3Dihub#APP1 , these fall-off thresholds require some assumptions to be met. E.g. FSC is between two independent half-maps; FSC vs resolution curve is a ‘nice shape’ - a threshold shouldn’t replace looking at the curve, which should be high before a single fall-off without big oscillations.

Whereas a half-map FSC for high-res cryo is trying to determine a threshold where correlated signal can be interpreted as scattering by the molecules, negative stain is giving you a lot of signal from scattering by variable stain envelopes.

Another quality measure could be to look at your map’s anisotropy. Relion for example automatically produces a 3D histogram of assigned particle angles, and Dmitry Lyumkis has shared his group’s program for measuring anisotropy with FSC: https://mail.ncmir.ucsd.edu/pipermail/3dem/2017-September/005210.html

Finally, you might want to know if the shape of your 3D map at low-res is correct, which is definitely not a given. You can collect tilt-pairs (or do tomography): https://www.sciencedirect.com/science/article/pii/S0959440X15000925 but I think there have been other validation methods suggested that don’t require extra data, but I can’t find the paper. T_T Does anyone know?

On 8 Sep 2018, at 01:00, Dmitry Lyumkis <[log in to unmask]> wrote:

Weekend FSC spar 2.0?

On Sep 7, 2018, at 11:34 AM, Chiara Rapisarda <[log in to unmask]> wrote:

Dear community members,

I would like to know what is the state of the art for reporting negative stain reconstruction resolution.

Is it best to not include it or to use the 0.5 FSC cut off. I definitely don't want to use the 0.143 cut-off, but I thought that it is important to give an idea of the quality of the reconstruciton by using some form of quantitative measure.

Is there an agreement on what to do?

Thank you for any feedback I will receive.

Chiara Rapisarda

Post doc
Université de Bordeaux, CNRS, INSERM
[log in to unmask]
Tel. +33 (5) 4000 3617

Institut européen de chimie et biologie
2, rue Robert Escarpit
33607 Pessac, France
www.iecb.u-bordeaux.fr

To unsubscribe from the CCPEM list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCPEM&A=1

To unsubscribe from the CCPEM list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCPEM&A=1

The Francis Crick Institute Limited is a registered charity in England and Wales no. 1140062 and a company registered in England and Wales no. 06885462, with its registered office at 1 Midland Road London NW1 1AT