[Demelerlab] AUC Collaboration: Sizing Amyloid Beta derived Oligomers

Tue Jul 25 14:55:42 MDT 2023

I am wholly confused about the y axis scale and label on your plot. It says
10 mm absorbance (i.e., 1 cm cuvette?). You should get values in the
0.0-1.0 range, not in the hundreds of OD. Assuming all of these
measurements are within the dynamic range of the detector, we should be
able to use any wavelength between 210-240 nm. We will adjust the
wavelength so that any concentration will absorb within 0.3-0.7 OD in a 3
mm centerpiece. Good to know that water+TFA against water does not
absorb at all, thanks for measuring this. It would still help us to know
what the actual scale on the y axis is. Can you send a corrected plot?

Thanks! -Borries

On Tue, Jul 25, 2023 at 1:36 PM Jason Zhu <jasonz13 at uci.edu> wrote:

> I reran the UV-vis spectra of water + 0.1% TFA, 1 mM F19Cha, 3 mM F19Cha,
> and 9 mM F19Cha all blanked against water and it worked much better this
> time. The TFA absorbs negligibly above ~220 nm and absorbs weakly compared
> to that of F19Cha. To get a 0.6 absorbance in a 3 mm centerpiece for the 1,
> 3, and 9 mM samples with less than 0.3 background absorbance, I would
> recommend 268, 278, and 279 nm respectively. I'm open to adjusting the
> wavelengths as you see fit. I've attached the Excel for the spectra below.
> I think the reason why I got negative absorbance last time was because the
> nanopure water that I was blanking with had some quality control issues.
>
> Please feel free to reach out if there's anything you need from my end!
>
> Best,
> Jason
> [image: image.png]
>
> Best,
> Jason
>
> On Fri, Jul 21, 2023 at 10:47 AM James S. Nowick <jsnowick at uci.edu> wrote:
>
>> Wonderful! Thanks, Bruce!
>> James
>>
>> On Jul 21, 2023, at 10:21 AM, Bowler, Bruce <Bruce.Bowler at mso.umt.edu>
>> wrote:
>>
>> Hi Borries,
>>
>> The samples arrived about an hour ago and are in my -20 freezer.
>>
>> I will mostly be around my lab until about 2:45 pm today.
>>
>> Cheers,
>> Bruce
>>
>> *From:* Borries Demeler <demeler at gmail.com>
>> *Sent:* Wednesday, July 19, 2023 9:26 PM
>> *To:* Jason Zhu <jasonz13 at uci.edu>
>> *Cc:* James S. Nowick <jsnowick at uci.edu>; Henrickson, Amy <
>> amy.henrickson at uleth.ca>; demelerlab at biophysics.uleth.ca;
>> maduni.ranasinghe at uleth.ca; Bowler, Bruce <Bruce.Bowler at mso.umt.edu>;
>> Frederick, Ariel <ariel.frederick at umconnect.umt.edu>
>> *Subject:* Re: AUC Collaboration: Sizing Amyloid Beta derived Oligomers
>>
>> Perfect! Thanks so much, and thanks to Bruce for handling our samples, I
>> will check in with you during the morning session of our workshop. Amy,
>> please remind me in case I forget.
>> Thanks, -Borries
>>
>> On Wed, Jul 19, 2023 at 9:24 PM Jason Zhu <jasonz13 at uci.edu> wrote:
>>
>> Hi Bruce, Ariel, and Borries,
>>
>> The peptide (F19Cha) should ship tomorrow (Thursday) morning and arrive
>> before 10:30 am on Friday. I've attached the shipping label and
>> instructions for how much water to add to make 1, 3, and 9 mM samples.
>>
>> Best,
>> Jason
>>
>> On Wed, Jul 19, 2023 at 11:59 AM James S. Nowick <jsnowick at uci.edu>
>> wrote:
>>
>> Hi All,
>>
>> The bottom line is that the absorbance of trifluoroacetate should be
>> negligible with respect to the peptide. The peptide absorbs so strongly at
>> 214 nm that you will need to go to longer wavelengths. Alternatively, I
>> think interference optics might be a good option.
>>
>> Best,
>> James
>>
>>
>>
>> On Jul 18, 2023, at 7:57 PM, Jason Zhu <jasonz13 at uci.edu> wrote:
>>
>> Hi Borries and James,
>>
>> I tried running different TFA concentrations on the nanodrop and got very
>> noisy UV-Vis spectra that I don't fully trust. I enabled the automatic
>> baseline correction and automatic pathlength adjustment settings on the
>> nanodrop which seems to have helped with the negative absorbance values
>> that don't make physical sense. I will meet with James tomorrow morning and
>> will work out the details of getting accurate absorbance values for both
>> TFA and F19Cha then.
>>
>> <image.png>
>>
>> I plan on weighing out the peptides, labeling them, and shipping them by
>> the end of the day Wednesday (tomorrow). I'll send an update once we figure
>> out the ideal wavelength for the different concentrations we plan to run at
>> each concentration and when the peptide is on its way.
>>
>> For lyophilized powder, 4C should be adequate. F19Cha isn't prone to
>> oxidizing from my experience working with it.
>>
>> Best,
>> Jason
>>
>> On Tue, Jul 18, 2023 at 2:38 PM Borries Demeler <demeler at gmail.com>
>> wrote:
>>
>> Hi Bruce and Ariel:
>> We would like to have some samples shipped to U of Montana, and I would
>> like to pick them up on Friday, if this is possible? The samples are
>> lyophilized peptide samples and can probably be stored at 4C. I will be in
>> Missoula on Friday  and Saturday, all day, our entire lab will actually be
>> in Missoula for a vaccine development workshop with the CTM group. Thanks
>> for helping again with the samples!!
>>
>> James and Jason:
>> Regarding the sample shipment, you can overnight them as a
>> lyophilized powder to:
>>
>> University of Montana
>> Dept. of Chemistry &Biochemistry
>> c/o Bruce Bowler or Ariel Frederick
>> 23 Campus Drive
>> Missoula, Montana 59812
>>
>> Please weigh them out and indicate on your shipping list with what volume
>> they need to be suspended to get the desired concentrations. Our loading
>> volume will be 110 ul, and we would like to know the wavelength to use
>> where we would get 0.6 OD in a 3 mm pathlength centerpiece, with no more
>> than 0.3 OD of background absorbance from the buffer (when blanked against
>> water). We can use different wavelengths for each sample to optimize the
>> absorbance for each concentration.
>>
>> The speed will be 60 krpm, so they should be measured in the An60Ti
>> rotor, at 20C.
>>
>> Please ship so they will be available for pickup by us on Friday this
>> week. Please indicate the storage conditions - I assume for the lyophilized
>> powder 4C is adequate?
>>
>> Thanks, -Borries
>>
>>
>> James S. Nowick
>> Distinguished Professor
>> Department of Chemistry & Department of Pharmaceutical Sciences
>>
>> Department of Chemistry
>> 4126 Natural Sciences 1
>> University of California, Irvine
>> Irvine, CA 92697-2025
>>
>> Phone:  (949)  824-6091
>> e-mail: jsnowick at uci.edu
>> Faculty Web Page: http://tinyurl.com/jsnowick/
>> <https://urldefense.com/v3/__http://tinyurl.com/jsnowick/__;!!CzAuKJ42GuquVTTmVmPViYEvSg!N1pni7MvQC2b2B8UkuEuyJfTSy3SNOX8O3u82A0rJS42kQP5CMvnPCF331ntQaU-5VV5CG3ejXgg-gw0D2sQwB0-qA$>
>> Research Group Web Page: http://tinyurl.com/nowickgroup/
>> <https://urldefense.com/v3/__http://tinyurl.com/nowickgroup/__;!!CzAuKJ42GuquVTTmVmPViYEvSg!N1pni7MvQC2b2B8UkuEuyJfTSy3SNOX8O3u82A0rJS42kQP5CMvnPCF331ntQaU-5VV5CG3ejXgg-gw0D2ux34kS7g$>
>> Pronouns: he/him/his
>>
>>
>> James S. Nowick
>> Distinguished Professor
>> Department of Chemistry & Department of Pharmaceutical Sciences
>>
>> Department of Chemistry
>> 4126 Natural Sciences 1
>> University of California, Irvine
>> Irvine, CA 92697-2025
>>
>> Phone:  (949)  824-6091
>> e-mail: jsnowick at uci.edu
>> Faculty Web Page: http://tinyurl.com/jsnowick/
>> Research Group Web Page: http://tinyurl.com/nowickgroup/
>> Pronouns: he/him/his
>>
>>
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