HT-SAXS Mail-In Instructions the basics

Introduction    Registration    Samples    Spreadsheet    Shipping    Data    Resources    Citing   

Introduction mail-in HT-SAXS at SIBYLS

HT-SAXS workflow

Below is a schematic diagram of the High Throughput Small Angle X-ray Scattering (HT-SAXS) workflow that you may take advantage of at the SIBYLS beamline 12.3.1 at the Advanced Light Source in Berkeley:


Mail-in-flow-june7

Robotics and detector

To achieve sufficient X-ray flux for informative scattering with low protein concentration and small volumes, we designed the SIBYLS beamline at the Advanced Light Source. We employ a light path generated by a super-bend magnet to provide a 1012 photons/sec flux (1 Å wavelength). The tunable incident wavelength enables rapid adjustment of the q range appropriate for the experiment without changing the sample to detector configuration (q=4π sin(θ /2)/λ where θ is the scattering angle and λ is the wavelength).

To make SIBYLS truly high-throughput, we installed a new Pilatus3 2M pixel array detector from Dectris, which gives much faster readout times over our previous MAR165 area detector.


Tecan_picture

SIBYLS then installed a new Tecan Freedom Evo liquid handling robot that divides the time it takes to collect a full plate in half, from approximately 5 hours to 2.5 hours per 96-well plate (with the potential to collect a plate in 10 minutes over the coming year).

We believe this will be a game changer for many reasons. We will be able to accommodate more users per week, enable new screening methods, and have more time to offer SEC-coupled SAXS and time resolved SAXS.

In addition to the new robotics, we have improved instruments for higher data quality. Our new monitor of transmitted beam intensity should greatly reduce buffer subtraction errors. We have also replaced the windows for lower background. In completing these upgrades, the stage is now set for increased sample to detector distance providing access to lower q (an upgrade that will occur sometime during the summer).


Registration request beamtime in 4 easy steps

1. Request mail-in account

All SIBYLS mail-in SAXS users must first register for a mail-in account:

This action is performed on this website. If you have never registered or are not logged in there will be a request link in the upper right hand corner of the page. You may also click the button below to get started.

Request Mail-in Account

2. Register with ALS

All users must also register with the ALS User Office:

You will not have access to your data until you are registered.

Register with ALS

3. Submit RAPIDD Proposal

Users must also submit one ALS RAPIDD proposal for every three mail-in uses:

Make sure you submit a "RAPIDD" proposal and not a "General User" proposal. RAPPID proposals are for record keeping only and will not affect your chances of using our service. So far, the high-throughput capabilities of our endstation have allowed us to accept every user that has requested mail-in SAXS time.

RAPIDD Proposals

4. Book mail-in slot

After requesting a mail-in account, registering with the ALS User Office and submitting a RAPIDD proposal, you may book a mail-in slot for beamtime:

If you are logged in to this site there will be a link to book mail-in time in the upper right hand corner.

In addition, the button below will lead you to the login or the mail-in booking page depending on your login status.

Login

Samples information on plate and sample requirements

Prepare 96-well plate

Order-plate-example

Our sampling robot uses specific 96-well Full-skirt plates. You must use this particular plate for your samples to be collected. Here is a link to order plates from Fisher Scientific: Order Plates


Order-cover-example

You will also need to use the AxyMat Sealing Mat for 96-well PCR Microplates to minimize evaporation. Here is a link to order mats: Order Mats


Sample requirements

Fill plate wells with 30µL of sample and buffer. Please keep the volume consistent throughout the plate.

Three sample concentrations between 1-10 mg/ml.

For redundancy, it is recommended to collect two (identical) buffers for each concentration series, one before and one after.

Do not use greater than 2% glycerol. It has been shown that 2% provides the maximum benefit. With glycerol, the viscosity of the solution increases and bubbles can become a problem. See our Methods paper for more information.




Spreadsheet sample entry and template

Upload spreadsheet

Our robot uses a spreadsheet to determine how to run your samples. Please follow the next four steps to stage your samples for data collection:

1. Download the spreadsheet template by clicking the button below:


Download Template

2. Fill it out as follows:

  • Column A

    ContainerID. "LabName". Write the name of your lab name here (no spaces).

  • Column B

    ContainerType. "plate". Do not change this column.

  • Column C

    Well. Indicates well location. Do not change any value in this column.

  • Column D

    SampleID. Write the name for the data output file (Sample ID). Only letters, numbers and underscores are accepted (no spaces).

  • Column E

    Collect. When checked ("x"), the sample will be collected (in order, starting from the top). If there is no sample or buffer in the well, leave it blank.

  • Column F

    Directory. Each sample will have it's own directory named after the well number. For example "A1", "A2", etc. If there is buffer in the well, add a "b" after the well number. This is very important for accurate data processing.

  • Column G

    WashAfter. Indicates a wash step. When checked, the sample cell will be thoroughly rinsed three times after that well is exposed. It is recommended to wash after a concentration series.

A B C D E F G
ContainerID ContainerType Well SampleID Collect Directory WashAfter
LabName plate A1 buffer1 x A1b
LabName plate A2 protein_1_low x A2
LabName plate A3 protein_1_medium x A3
LabName plate A4 protein_1_high x A4 x
LabName plate A5 buffer1 x A5b
LabName plate A6 buffer2 x A6b
LabName plate A7 protein_2_low x A7
LabName plate A8 protein_2_medium x A8
LabName plate A9 protein_2_high x A9 x
LabName plate A10 buffer2 x A10b
LabName plate A11 buffer3 x A11b
LabName plate A12 protein_3_low x A12
LabName plate B1 protein_3_medium x B1
LabName plate B2 protein_3_high x B2 x
LabName plate B3 buffer3 x B3b
LabName plate B4 B4 (no sample)

3. Upload your updated spreadheet to the SIBYLS SAXS database

Upload Spreadsheet

4. Once you have booked a mail-in slot, a unique barcode will be created for your plate. Print this label out and tape it to the side of your plate.

Example Label:

Barcode_image_example