The LPS I used was bought in SIGMA L6261 (http://www.sigmaaldrich.com/catalog/product/sigma/l6261?lang=pl®ion=PL). I dissolved in PBS in 1mL (1mg/mL) and it worked very well. I added 100uL of LPS diluted in PBS to each eppendorf to avoid freeze/defreeze. According to manufacturer's instruction you can store it at -20 C for 2 years and between 2 - 8 C for 1 month.

If you need something, do not hesitate to ask me.

Proteins in general should be stored in buffered solutions (phosphate, TRIS, HEPES, etc). 150mM NaCl also can help. pH 7-8 is usually good, but will vary depending on the protein. If pure protein can be stored at 4C for up to a week, but again this depends on the propensity for the protein to degrade. As for freezing and thawing a buffered solution with salt should be fine. If you find that the protein crashes out upon thawing you can try freezing with glycerol (1-10%v/v). Determining the best condition to freeze and thaw usually takes for experimenting. Also lower conc are less likely to aggregate when thawed and freezing quickly with dry ice can also be beneficial.

LPS is a polysaccharides. we usually prepare a fresh solution everytime.

If you want to use LPS with a narrow variance you should not store diluted samples. LPS adsorbs to many surfaces. Consequently, you lose high proportions at low concentrations without knowing the actual amount. It is also mandatory to use pyrogen-free water for dilution, e.g. water that is supplied in ampoules for single use and certified for LPS absence.

In my case. I diluted LPS in DMEM media, stored at 2-8C, and used within a weak. But I think the best way is using diluted solution immediately after prepared

We store our Ultra-Pure LPS @ 10mg/ml stock solutions in sterile-filtered PBS, pH 7.2. @ volumes of 10µl/200µl tube at -20oC. Sterile-filtration is done on the PBS, not the resuspended LPS. Each small tube is thawed at most 2 times before discarding. Thawed LPS is diluted in the solution of interest, usually cell culture media, as a sub-stock and vigorously vortexed for 1-2 minutes. This sub-stock can then be added to the media/sample of interest. In this way we keep the stocks concentrated and minimize thaw cycles.

Oh, sorry. We usually use up these samples within 6-9 months. Longer than a year it's not so good.

Good luck!

Normally, I store 1mg/mL of LPS in DMEM media or non-phenol red media in -20 degree celcius. Based on my experience, diluted LPS cannot be stored for long period of time. Diluted LPS is best to use immediately after prepared.

Thanks for help everybody!

The LPS I used was bought in SIGMA L6261 (http://www.sigmaaldrich.com/catalog/product/sigma/l6261?lang=pl®ion=PL). I dissolved in PBS in 1mL (1mg/mL) and it worked very well. I added 100uL of LPS diluted in PBS to each eppendorf to avoid freeze/defreeze. According to manufacturer's instruction you can store it at -20 C for 2 years and between 2 - 8 C for 1 month.

If you need something, do not hesitate to ask me.

Thanks again!

We followed exactly the same steps Juan, and Yes its works. We treted human PBMCs with LPS from E. colli (SiGMA). Just instead of PBS we used NaCl (0,9%)/saline1

I am  naive in the use of LPS. I am using it to challenge the immune system of bats and then recored the energetic costs associated withe mounting the immune response. I using the same protocol as described by Dr. Juan Antonio Giménez Bastida to prepare and store the samples. However I stored it at 2-5C and the samples had been in this condition for 6 months now. I am doing some pilots now and, much to my surprise, bats are not responding to injections of these solutions (so far I did 50 and 100ul injections). I am wondering if the absence of the response is not due too the fact that the solutions were not properly stored. 

Hi Ariovaldo,

You should check the storage conditions described in the manufacturer's sheet. See the stability of your solutions at 2-5 C. If the solution you prepared is stable for 1 month in these conditions, it could be explain why your experiments do not work.

You also can consider that the concentration used is not enough to induce any response in the animals.

Hope that helps,

If you have any question, do not hesitate in asking me,

Best,

Juan.

Hi Juan,

Should we use 1xPBS or 10XPBS?  In addition, we purchases 10XPBS from Bio-rad.

Thanks in advance,

Best,

Feng

Hi Feng Li,

I always use 1xPBS.

Regards,

Juan.

Thanks, Juan.

Feng

Can I dissolve 25mg LPS powder in 1 ml PBS and make the aliquots ?

yes you can do this, no problem. 

Thanks, Ariovaldo : )

Jiang

Hi Dr. Juan Antonio Giménez Bastida.

I want ask base on your experience in the use of LPS so far, do you have any recommendable the temperature at which you've been storing LPS in your laboratory and still maintain potency?

Thanks.

Hi Olatunji,

I always follow the recommendations given by the manufacturer. I like keeping at -20C, although I also have used LPS for 6 months kept at 2-8C and it maintained its potency. 

Best,

Juan.

Hello Praveen Jaiswar,

I have never kept LPS at -80 C. Therefore, I do not know if it can be degraded or damaged. I always keep it at -20 C following the manufacturer's instructions.

Hi Oskan Tasinov,

I agree with Juan Antonio Giménez Bastida. I also had LPS from Sigma and you do not have to kept LPS at -80 C. On my PhD studies, when i was using LPS to treat me cells i dissolved  LPS in PBS in 1mL (1mg/mL). I was also adding 100uL of LPS diluted in PBS to each eppendorf to avoid freeze/defreeze. 

If i can help in something also then please ask ::

Thank you all for your suggestions! It really helped me, I already defended and know helping my colleaques to do the same experiments. I observed when they tries to change the treatment concentrations very high (close to 1mg/ml) and to use the some aliquots within 1 year (less than 2) they can still give different (usually decreased) immune responce induction potential.

Dear colleaques,

I hope this discussion was useful for every one, good luck in your projects!