156 replies to this topic

[resveratrol_guy]

 

Another update from TLR.  

 

Hi Mischa,

After receiving what was by far the best quote for rhDNase1 and remitting the offer to you, bearing I knew it was still likely higher than what was feasible, I subsequently inquired to a colleague of mine who specializes in protein expression optimization as to his thought to creating an optimized system and within that what best cost basis could be garnered. Usually he gets around $4000 upfront for this service, but as we both do many favors for each other and that he strongly supports Project TLR he was willing to explore it at no charge. 

Within this, his initial assessment is that an optimized expression system that he can derive should be able to provide a cost basis that is about half of the prior pricing offered. 

 

If something of that nature is workable for you I will inform him to proceed. At this point he does not know the actual firm cost basis with certainty, but he seems confident within pursuing it he can get the expression to yield means for production that produces the ability to offer a price that is around that price basis.

 

So, let me know if this seems to be within the area that will be viable.  Such would apparently seem to have ability to be under $1/mg basis, which I hope is tenable.  If so, he will than work to refine the expression and look to achieve a best cost basis that should hopefully be somewhere in accord with this initial estimated cost basis.  Once such is established, from there I will remit the firm offer to you based on such. 

Brgds~

 

 

The price at the top of the previous page already equates to $0.4/mg, so "under $1/mg" would not necessarily be an improvement. That aside, if it's really half as much as above, then we're talking around $0.2/mg, or about $24/d at the study dose.

 

I would be in favor of pursuing this source, but we would need to commit to a competent third party lab to do the CoA. We would need a strong authentication protocol involving a homogenization of the mixture, which would detect scams involving real stuff on the top and fake stuff on the bottom. I would let them know up front that we're going to turn the stuff inside out looking for inpurities, such that we require 98% or 99% or whatever. There are plenty of labs out there who could assist.
 

Edited by resveratrol_guy, 22 July 2016 - 04:48 AM.

[deetown]

You're not concerned with the potential immune reaction from bovine Dnase?

 

For testing it seems the way to test this stuff is using a western blot test.  It's something any small lab does on their own requiring minimal equipment.  Thus It's been hard to find a lab to test it.  One quote from a lab for more complex testing came back at over $6k.  Team TLR said they would be fine with meeting someone in person doing the blot test together before the transaction.  

Edited by deetown, 22 July 2016 - 05:17 AM.

[Mian Ali Ismail]

I have heard there is a supplement of the bovine form available in Russia ?Can anyone find the supplement company? I tried but failed.

[resveratrol_guy]

deetown, yes, I am concerned about an allergic reaction to the bovine form, and I would certainly prefer the human form. Unfortunately, I'm not seeing a lot of options here.

 

Ali, I really wouldn't want to obtain anything from Russia. The scam rate is very high.

 

I hate to say this, but I think we've pretty much exhausted the options that are even somewhat promising. Even if it were possible to obtain genuine Pulmozyme without a prescription, the price ends up being like $100+ per mg, so that's like $10,000/day. If it cures Alzheimer's in a week, that would probably be worthwhile, but I think if it works at all, it will need to be taken periodically, perhaps a few times per month.

 

TeamTLR appears to be "the" choice. So unless anyone else has a serious alternative, I suggest we move forward to obtain a firm quote from them. We're not getting any younger.

[deetown]

Logic was waiting to hear back from a few labs I believe.  But assuming this is the best option let's see how much interest there is.  So assuming testing costs $1000 for travel expense and testing materials.  The price would be around $430 per gram.  I'd be in for 5 grams.  Who else is interested at that price and at what quantity?

 

Edited by deetown, 23 July 2016 - 05:44 AM.

[plumper76]

I could do 2 grams at that price for now...

[stefan_001]

I ran into this page, collection of dnase1 manufacturers and its uses. Seems its quite often used in lab experiments.

http://www.labome.co...nt/DNase-I.html

[stefan_001]

We have previously reported that the urine of febrile humans contained large quantities of an inhibitor of IL-1-induced murine thymocyte proliferation that was a glycoprotein between 30 and 40 kD in size. In the present study this factor has been purified to homogeneity using a sequence of eight purification steps (ammonium sulfate precipitation, ion exchange chromatography, molecular sieve chromatography, hydrophobic affinity chromatography, hydroxylapatite chromatography, fast protein liquid chromatography, and two HPLC steps). SDS-PAGE analysis indicates that the purified material is a 38-kD molecule. Evidence based on a partial amino acid sequence analysis as well as enzyme studies indicates that this inhibitor is a type of human DNase I.

https://www.ncbi.nlm...les/PMC2189114/

[stefan_001]

So here a wild suggestion. From this study:

 

The results reported in this paper show that the pancreas could be a tissue of origin of the major electrophoretic form of endoDNase type I activity of human serum

http://www.jbc.org/c.../24/12588.short

 

So increasing the pancreas secretion would lead to serum level DNASE increase? Eat "food" every hour to keep it working? Perhaps that can be an alternative route?

 

Using sham feeding in humans, one study [113] demonstrated that sham feeding stimulated pancreatic enzyme secretion at up to 50% of the maximal secretory rate with no increase in bicarbonate secretion when gastric secretions were prevented from entering the duodenum. When gastric secretions were allowed entry into the duodenum, the rate of pancreatic enzyme secretion rose to about 90% of the maximal. 

http://www.ncbi.nlm....books/NBK54132/

Edited by stefan_001, 24 July 2016 - 07:55 PM.

[resveratrol_guy]

We have previously reported that the urine of febrile humans contained large quantities of an inhibitor of IL-1-induced murine thymocyte proliferation that was a glycoprotein between 30 and 40 kD in size. In the present study this factor has been purified to homogeneity using a sequence of eight purification steps (ammonium sulfate precipitation, ion exchange chromatography, molecular sieve chromatography, hydrophobic affinity chromatography, hydroxylapatite chromatography, fast protein liquid chromatography, and two HPLC steps). SDS-PAGE analysis indicates that the purified material is a 38-kD molecule. Evidence based on a partial amino acid sequence analysis as well as enzyme studies indicates that this inhibitor is a type of human DNase I.

https://www.ncbi.nlm...les/PMC2189114/

 

Is there any realistic chance that we could use this technique to create human DNase 1, even assuming that the paper is valid? I think the probability is low, considering that the input is urine, which is rich in all sorts of compounds we don't want.

 

I'm still undecided on how much if any TeamTLR product I would want to buy. I might go for a gram if there was not enough interest otherwise.

[Mian Ali Ismail]

The researcher hinted towards a bovine form of Dnase 1 available i Kazakastan as a supplement.We could also get the bovine form from sigma aldrich which is relatively a lot cheaper.And he said it should be taken in capsule form otherwise the enzyme would get destroyed in the stomach.

[deetown]

Resveratrol Guy, I haven't had any luck with a lab.  Let me know if you know of one.  

[resveratrol_guy]

Honestly I don't think this is going anywhere. I'm not even worried about a lab, in the absence of a viable source.

 

Ali, do you have any further details on the Kazakhstan supplement? Maybe a brand name or manufacturer? (Not that I'm really optimistic here.)

[Mian Ali Ismail]

Honestly I don't think this is going anywhere. I'm not even worried about a lab, in the absence of a viable source.

 

Ali, do you have any further details on the Kazakhstan supplement? Maybe a brand name or manufacturer? (Not that I'm really optimistic here.)

 I asked the researcher he said the manufacturer stopped making it due to low demand.He also told the supplement was bovine form and sigma aldrich also gives bovine form Dnase 1 which is a trustworthy source I think.

Can this also be used for neurodegenerative disease ? 

 

http://www.medgadget...oming-coma.html

[ceridwen]

I'm suspicious about anything bovine due to further prion disease risk!

[resveratrol_guy]

 

Honestly I don't think this is going anywhere. I'm not even worried about a lab, in the absence of a viable source.

 

Ali, do you have any further details on the Kazakhstan supplement? Maybe a brand name or manufacturer? (Not that I'm really optimistic here.)

 I asked the researcher he said the manufacturer stopped making it due to low demand.He also told the supplement was bovine form and sigma aldrich also gives bovine form Dnase 1 which is a trustworthy source I think.

Can this also be used for neurodegenerative disease ? 

 

http://www.medgadget...oming-coma.html

 

 

Yeah Sigma Aldrich makes trustworthy products, which are impossible to acquire in the US but might be easily acquired in places such as Kazakhstan. Too bad they stopped making it.
 

[Mian Ali Ismail]

IF the supplement might have been available it would have given us a fair chance 

[Logic]

I  have just sent of another Email to those companies that replied.
I don't hold out much hope for this.  :(

[Logic]

Good News! Possibly...

 

Manufacturer:

 

"...Unfortunate news; the project did not result in a confirmative response. The rhDNase may not be suitable for a catalogue protein. The labs are going to try again if possible, but we may need to do so as a custom order instead of waiting for it to become a catalogue item.

I thank you for your patience through all of this, and I wish I could report with better news. I do hope you may still be interested in trying further projects with us, but either way I do appreciate your interest..."

 

My reply:

 

"...Thanks for all your effort in this.
It would seem that we are now at the difficult point where a the cost for a custom order/synth depends on the size of the order, and the size of the order depends on its cost. 
I will pass on our correspondence to the interested parties and get a ballpark order size that way.
I will then get back to you for a quote..."

 

So; assuming a reasonable price, how much Dnase do we want!?

Edited by Logic, 21 September 2016 - 02:13 AM.

[psychejunkie]

Exercise is a potent stimulus for enhancing circulating DNase activity

 

 

Results

Serum cfDNA concentration was significantly (P ≤ .001) elevated immediately post (2.5-fold) and 7 min post exercise (2.3-fold) with a return close to baseline at 30 min post exercise (1.5-fold). The rise in cfDNA was accompanied by a concomitant, significant (P ≤ .001) decrease in serum DNase-AR from 15.1% prior to exercise to 3.1% AR at cessation of the exercise test and 7 min post exercise (3.9% AR). DNase-AR returned close to baseline at 30 min post exercise (5.2% AR).

Conclusions

A single bout of high intensity exercise is a potent stimulus for enhancing circulating DNase activity in healthy people. Acute exercise may therefore be considered as a non-pharmacological stimulus to trigger DNase activity.

...

source: http://www.sciencedi...009912013006024

 

[Der Springende Punkt]

Good News! Possibly...

 

Manufacturer:

 

"...Unfortunate news; the project did not result in a confirmative response. The rhDNase may not be suitable for a catalogue protein. The labs are going to try again if possible, but we may need to do so as a custom order instead of waiting for it to become a catalogue item.

I thank you for your patience through all of this, and I wish I could report with better news. I do hope you may still be interested in trying further projects with us, but either way I do appreciate your interest..."

 

My reply:

 

"...Thanks for all your effort in this.
It would seem that we are now at the difficult point where a the cost for a custom order/synth depends on the size of the order, and the size of the order depends on its cost. 
I will pass on our correspondence to the interested parties and get a ballpark order size that way.
I will then get back to you for a quote..."

 

So; assuming a reasonable price, how much Dnase do we want!?

 

Great, Logic! 120 mg would be a daily dose. 2 months should be enough to see results so 7 g could be an estimated amount for a single person.
 

[Logic]

 

Good News! Possibly...

 

Manufacturer:

 

"...Unfortunate news; the project did not result in a confirmative response. The rhDNase may not be suitable for a catalogue protein. The labs are going to try again if possible, but we may need to do so as a custom order instead of waiting for it to become a catalogue item.

I thank you for your patience through all of this, and I wish I could report with better news. I do hope you may still be interested in trying further projects with us, but either way I do appreciate your interest..."

 

My reply:

 

"...Thanks for all your effort in this.
It would seem that we are now at the difficult point where a the cost for a custom order/synth depends on the size of the order, and the size of the order depends on its cost. 
I will pass on our correspondence to the interested parties and get a ballpark order size that way.
I will then get back to you for a quote..."

 

So; assuming a reasonable price, how much Dnase do we want!?

 

Great, Logic! 120 mg would be a daily dose. 2 months should be enough to see results so 7 g could be an estimated amount for a single person.
 

 

 

Thx!  

I'll go back through the thread and thumb suck an amount based on the # of interested people X7 then shall I?

[Logic]

Ok; here is a list of people that have posted in this thread:

 

1:   Mian Ali Ismail
2:   resveratrol_guy
3:   deetown
4:   gamesguru
5:   Alex_G
6:   Der Springende Punkt
7:   Linda Gray
8:   plumper76
9:   45rpm

10: LongLife
11: psychejunkie
12: davis89x
13: Jochen
14: centralFloridaMan
15: mrwhitee
16: MarcB
17: Major Legend
18: stefan_001
19: ceridwen
20: psychejunkie

 

From my experience in the Nilotinib group buys I am going to assume that around 70% = 15 people will join the buy.

That's 15 X 7 grams = 105 grams..?

 

I have sent an invite to the above list.

Plz post your interest and the amount of Dnase you would be interested in assuming a reasonable price so that I can get a quote.

 

[ceridwen]

Interested thanks

[deetown]

20 grams 

[Der Springende Punkt]

In for 7 grams.

[resveratrol_guy]

7 grams

[plumper76]

In for 7 grams

[TRUGAN]

possibly 7 grams ...depending on price.

[TRUGAN]

hmmm... Sorry as I havent kept up with the thread but is this a bovine source?