Sodium Bicarbonate - Science topic

Hi, could you please explain to me how to prepare 500 mL of coupling buffer to CNBr activated Sepharose Beads containing:

-0.2 M Sodium bicarbonate

-0.5 M of NaCl

Final volume= 500 mL; pH=8.4 (adjust pH with NaOH).

Thank you.

Dear all,

I have just purchased an aliquot of lncap from atcc, I thawed them and the result was disastrous. Briefly, these cells form big clusters and even when they adhere they maintain this morphology which seems far from the one in literature. Atcc told me that the RPMI I used (which is the same I employ for other lines) is too low in glucose, it omits sodium pyruvate and has HEPEs and sodium bicarbonate different from the one in the suggested RPMI. My ignorance, I wasn't aware that differences between RPMI from different suppliers were so important. In addition they told me I made a mistake using heat inactivated FBS as, probably, I eliminated some growth factors.

Could you kindly tell me if, in your opinion, are these requirements all mandatory for a satisfactory culture...it is so crazy. Thank you in advance

I have degummed the silkworm cocoons with 0.03M sodium bicarbonate and dried it overnight at room temperature, and added 9.3M LiBr on the top of dried fibers in the 20:80 ratio but my solution becomes gel instead of dissolving.

What water should be used in a water jacketed CO2 incubator? The manual says to use "sterile distilled water", and when asking VWR by telephone, or looking at ThermoFisher online, they say it should be 50K - 1 M Ohm, and pH 7-9. We have a MilliQ machine for ultrapure water, nevertheless we ordered Poland Spring distilled water since the ultrapure is 18 M Ohm. However, I looked up the specs, and the Poland water is pH5. Calling VWR they didn't resolve it. They want the given parameters, and he worries about the effect of adding something like sodium bicarbonate.

Is everyone else simply not worrying about this? What do you use?

I have a BSA-FITC conjugate made in Sodium Bicarbonate pH 8.5 that I'd like to desalt (the excess dye) and buffer exchange (into PBS) via gravity. I equilibrated a G-25 column with PBS pH 7.4. I applied my conjugate (2.5 mL volume) and eluted it with 3.5 mL PBS so my fraction 2 (2.5 mL volume) is my conjugate in the correct buffer confirmed by pH and OD at 280 nm. What I don't understand is my next fraction F3 (3.5 mL volume) has a pH of 8.5, the bicarbonate buffer, but also has protein (maybe?). OD says there's no protein detected, but by agitating this fraction there seems to be protein in there. Also, by gravity the conjugate should be diluted, but my F2 has the same volume. It sounds to me I want to combine my F2 and F3 (hence why after desalting, sample comes out more diluted), but my F3 now has sodium bicarbonate buffer?

I'd like a powdered form of MCDB 131 Medium with sodium bicarbonate added, and I've noticed that not only is the powder form of this medium difficult to find with NaHCO3 already included, but the same is true for other media like DMEMs. Liquid versions that include NaHCO3 are plentiful. So are powders made this way because the amount of CO2 exposure varies between cell lines/projects and the concentration of buffer therefore needs to vary as well? Or is there some concern with NaHCO3 interacting with other components in its powdered form? Or perhaps it has to do the with manufacturing process for powders? I can't think of a reason that milling in an industrial mill with ceramic stones, for example, would be problematic for sodium bicarbonate, though. I'd love any insights on this.

I want to prepare mammalian cell culture grade Sodium bicarbonate(SBC) solution from powder available in media kitchen for regular use.

Can i make it this way?

1. Dissolving SBC powder.

2. Filtering by 0.22um filter

3. pH adjustment

4. Sterilization by autoclaving

Does autoclaving degrade SBC?

Does pH will change after autoclaving?

Resuscitation drugs become ineffective during severe metabolic acidosis ...why can't bicarbonate be a part of ACLS protocol

Resuscitation drugs become ineffective during severe metabolic acidosis ...why can't bicarbonate be a part of ACLS protocol

I am looking for answers as to whether NaHCO3 buffered cell culture media stored outside a CO2 gassed atmosphere changes it's pH irreversibly.

I understand how the NaHCO3 buffer principally works. But I wonder if the equilibrium between CO2 and HCO3- is reversible, always readjusting, depending on ambient temperature and CO2 gassing (+ atmospheric pressure)?

Suppose I would prepare a medium that is NaHCO3 buffered and has a pH = 7.3 after preparation in the lab (i.e., 20° C & 0.04% CO2). The pH was adjusted using HCL and NaOH. If I were to incubate this medium for 24 hours at 5% CO2, 37°C, and then remove it from the incubator, would the pH then return to 7.3 at the Lab-atmosphere?

This would be easy to prove experimentally and I will try this out in the next few days, but I would be interested to know if I might be missing something.

I am currently attempting to culture cell lines in a high %CO₂ incubator to mimic hypoxic conditions. Unfortunately we do not have incubators that can adjust the O₂, nor do I have access to a hypoxic chamber so increasing the CO₂ to 20% seems to be my only option.

The resulting issue: cell culture media typically contains a sodium bicarbonate buffering system that is optimised for incubators set between 5-10% CO₂, so in a 20% CO₂ incubator the media becomes slightly acidic.

Theoretically, I could increase in concentration of NaHCO₃ to 8g/L for 20% CO₂ to buffer the media to a pH of 7.4 (a reference for the calculation used to obtain this value https://www.researchgate.net/deref/https%3A%2F%2Ftools.thermofisher.com%2Fcontent%2Fsfs%2Fbrochures%2FD19558.pdf?_tp=eyJjb250ZXh0Ijp7ImZpcnN0UGFnZSI6InNpZ251cCIsInBhZ2UiOiJxdWVzdGlvbiIsInBvc2l0aW9uIjoicGFnZUNvbnRlbnQifX0), this however leads to changes in the osmolarity that my cell lines can't seem to handle.

Does anyone have any suggestions on how I could adjust my cell culture media to suitably culture cells in 20%CO₂?

I have recently purchased B16-F10 cell line from NCCS Pune. They sent me cells of passage 28. I am using DMEM, high glucose with L-glutamine and sodium pyruvate and 3.7 g/L sodium bicarbonate+10%FBS media in 5% CO2 incubator to culture these cells.

After culturing for few days, I am not able to see any melanin pigment in the cytoplasm. They are completely amelanotic. I need these cells to secrete this pigment. Can anyone help me in troubleshooting this?

I have been attempting to grow BT474. I have taken the cells out of cryogenic storage, but each time there is no attachment to the flask I have placed them in. I'm using RPMI 1640 media (with L-Glutamine and Sodium Bicarbonate) and with 20% FBS, I previously used 10% FBS, the increase has made no difference. Has anyone had a similar experience with this cell line? If so how did you solve this issue. Any suggestions are welcome.

I work in a single-molecule analysis lab. I have been waiting on a supplier to send sodium bicarbonate for a while, and I want to begin experiments now. We have a stock of sodium bicarbonate from 2017 (five years old). Why can't I use this? Does it really have an expiration date?

I imagine if it does, it may have to do with coming into contact with the air after it was opened so many years ago.

Any help is appreciated. I dont want to do all the experiments only to have them invalidated by the fact that I used old sodium bicarbonate.

Sodium bicarbonate is among five evidence-based performance supplements (caffeine, creatine, nitrate/beetroot juice, β-alanine, and bicarbonate). I need a pragmatic approach to the culinary use or the use as a supplement of this compound. NaHCO3 ingestion should be consumed at a dose of 0.2–0.4 g/kg BM. Moreover split doses taken over a 30- to 60-min time period or serial loading with three to four smaller doses per day for two to four consecutive days prior to an event has been proposed.

I want to prepare media for cell culture use. Hence, I need to get water sterilized and add sodium bicarbonate and sodium pyruvate to it post-sterilization. I tried a different method, where I added sodium pyruvate and sodium bicarbonate to water and kept it for autoclaving. later I was able to see precipitates in the solution. Can anyone suggest what chemical reaction might have taken place. I found few articles suggesting the release of CO2 from sodium bicarbonate by autoclaving. But the precipitate formation has not been mentioned anywhere. Please suggest me what can be the error from my end.

Can someone tell me if Dulbecco′s Modified Eagle′s Medium - high glucose With 4500 mg/L glucose, L-glutamine, sodium pyruvate, and sodium bicarbonate, liquid, sterile-filtered is suitable for CHO-K1 cell growth and in vitro toxicology research? Will the cells grow well in such a medium? Can the components of the medium affect the toxicity of the metals to the cells?

Thank you in advance for all answers.

I want to add sodium bicarbonate in RPMi.

I have sodium bicarbonate concentrate and it says 0.1M eluent concentrate for IC.

Can I use it at human cell culture?

How can I add sodium bicarbonate in RPMI?

I heard that by adding it, it will change pH..

adjust pH, autoclave, and add it to RPMI is right?

Please give me a help.

Thank you!

I am looking for an alternate method to measure percentage of carbonate impurity from sodium bicarbonate. The specification is <0.23% carbonate. There is a titration method using barium chloride to precipitate the carbonate and back titration for the bicarbonate. This will measure total carbonate/ bicarbonate and also bicarbonate only, in order to calculate the carbonate. I am not sure if this will be sensitive enough for small amount of carbonate, which is less than 0.23%. I try find other ideas or separation with different techniques, for example using ion chromatography, but it seemed that carbonate and bicarbonate not separable by IC by many articles. Any comment and suggested techniques or method for this quantitation. This was originally a project to find alternate method from the USP method using the carbonate apparatus (for sodium bicarbonate) since we have difficulty with the method.

Hello, I',m having quite some difficulty growing Ramos (EBV-negative Burkitt lymphoma) cells. I'm using a modified RPMI 1640 culture medium to match ATCC's recommendations (RPMI 1640 + 10% FBS + 25 mM HEPES + 2 mM L-glutamine + 1 mM sodium pyruvate + 4500 mg/L glucose + 1,500 mg/L sodium bicarbonate). It has been over a week since I resucitated a cryovial, and although the cells are not dying, they are not multiplying either as expected. I have changed medium every 2-3 days as suggested by ATCC.

Any suggestions on what to do to make them grow?

Thank you!

I am using TC-100 media(with serum and antibiotics) having sodium bicarbonate and L-Glutamine to grow Sf21 cells, but after I thaw them, they initially attach and grow for some time, then they fail to attach after 2-3 passages and eventually die.

What could be the possible reason for it? How can this be tackled?

Sample tube: crude enzyme+ 1 % casein (100mM sodium bicarbonate buffer pH-9) than incubate for 30 minute add TCA (5% in distil water) than centrifuge than take readings at 600 nm.

Blank1: water + casein, same as above

Blank 2: water + enzyme, same as above.

take readings from spectro. (Run Sample and Blank simultaneously)

I am working with MCF 7 cell line. I ordered the cell line from NCCS Pune. I received a very high confluent flask. After a day, I splitted the cells. I am using MEM w/Earle's salts, 2mM L-Glutamine, 1mM Sodium pyruvate, NEAA and 1.5 gms per litre Sodium bicarbonate with 10% FBS and antibiotics. A day after the splitting, I observed that all the cells were detached and floating. I changed the media and the flask again but next day the cells were in same condition ? They are not adhering in the flask and a lot of cells are dead. What should I do ?

Hi there,

I've been noticing that after adding my blocking chemicals (TEA-Cl and CdCl2) to isolate sodium current, my pH increases beyond my ideal point (I'm trying to stick around 7.4 and it drifts to around 8.0 after adding both) and there is first cloudyness and then precipitate forming in the solution after addition of the CdCl2.

Prior to adding the blockers, I am bubbling my ACSF with carbogen for twenty minutes. I also do not add sodium bicarbonate or dextrose to my 10X stock. My concentrations for ACSF (working solution) are as follows:

125mM NaCl

2.5mM KCl

1.25mM NaH2PO4

1mM MgCl2

25mM NaHCO3

25mM dextrose

2mM CaCl2

75mM TEA-Cl

0.2mM CdCl2

Again, target pH is 7.4. Would really like some input on this, as well any any relevant chemistry as to what is happening. Thank you!

Hi

I have Biosra brand DMEM high glucose powder medium, which I prepare according to the protocol. But after adding 3.7 g/L of sodium bicarbonate. I set the pH of the environment between 7.3 and 7.4. At first, the color and pH of the environment are good, but after a short time, the color becomes pink and the pH is alkaline. what should I do? Do I need to add HEPES? And if so, how much?

I’m working on HIT_T15 cells , and I would ask about MTT assay!!

i Know that the medium should be without phenol red , but most of the mediums I found it available was without phanol red and l_glutamine or sodium bicarbonate also or without HEPES , what should i do in this case ? May any one help me please

I’m working on HIT_T15 cells , and I would ask about MTT assay!!

i Know that the medium should be without phenol red , but most of the mediums I found it available was without phanol red and l_glutamine or sodium bicarbonate also or without HEPES , what should i do in this case ? May any one help me please

Hi I am growing marine algae however i have lots of contaminants in my samples such as rotifers . I have attempted to remove these with sodium bicarbonate however the eggs still do remain. i have tried filtration with a 0.45um filter however the filter cloggs. Any suggestions on how i can remove these without losing the algae as the aim is to have a pure culture?

I found that there are two pKa values of sodium bicarbonate i.e 6.4 and 10.3. So can I take sodium bicarbonate to prepare buffer solutions of pH values around 6.4 and 10.3? Please suggest me.

Is 0.2M sodium bicarbonate always has pH 8.3? I mean do you need to add HCl/NaOH to adjust the pH when you mix 0.2M sodium bicarbonate with water? Really confused about it.

Soda bicarb is considered an ideal buffering agent to maintain rumen pH. Rumen has a large volume. Different literatures give different answers. How can we arrive at an ideal oral dose rate, interval and period for this buffering action in a practically possible manner?

I have a body of water (volume=x) with about pH: 5.5 and alkalinity of about 4,5 mg/L CaCO3. I am looking to increase the pH to about 7,2 and increase the alkalinity as much as possible (no more than 200mg/L CaCO3). Is there a correct way to calculate the amount NaHCO3 needed for this? What other parameteres are needed for the calculation?

CO2: 5%

O2: 95%

Role of pH in RBC lysis solution

I want to ask aquestion in context of RBC lysis solution used in flow cytometry

We have been using our own lab made lysis solution containing ammonium chloride sodium bicarbonate and edta but when we keep the cells for more than 15-20mins in the solution out neutrophills convert into debris Or start to loose there integrity and shift to lower forward and side scatter.

Does pH play an important role here?

How long we can store the solution?

Hi.

I want to perform a plaque assay using MA104 cell line. After virus absorption, I wash the cells with serum free media and add a 1:1 mixture of 1.2% agarose and Medium( DMEM containing L-Glutamine, Vitamins, Nonessential amino acids, sodium pyruvate, sodium bicarbonate and Pen/Strep). I put the plate in 37C and every 24 hours I check them to avoid gel drying. I check them every 24 hours but after 48 hours, cells die and I can not see any plaques.

Does anyone know how can I keep them alive to continue the test and observe the plaques?

Recently Sigma and other companies are providing culture media that are modified for autoclaving. For example: Auto-Mod™, without L-glutamine and sodium bicarbonate, powder, suitable for cell culture (Sigma R7755). Do these autoclavable media work as well as sterile-filtered media?

Hi everyone, I am currently culturing NK-92 cells in the ATCC recommended culture medium (Alpha-MEM without ribonucleosides and deoxyribonucleosides but with 2 mM L-glutamine and 1.5 g/L sodium bicarbonate. With: 0.2 mM inositol; 0.1 mM 2-mercaptoethanol; 0.02 mM folic acid; 100-200 U/ml recombinant IL-2; 12.5% horse serum and 12.5% fetal bovine serum)

The culturing has been successful before for a few weeks however now when using a thawed out sample (from liquid nitrogen) I see under the microscope that the cells are not forming the proper (grape-shaped) clumps. In fact they are quite far from globular shaped and are very oddly-shaped. Has anyone had this issue before? My coworker is culturing the same thawed-out cells in the same medium but hers form proper clumps, while mine are very strangely shaped. Any suggestions or ideas would be appreciated, thank you!

I´m currently working with the THP-1 cellline.

I have two questions.

1. I got those cells as a gift from a nother group which told me to cultere this cellline in RPMI + Glutamin + Pen/Strep.

i recently read that they also need 10 mM HEPES, 1 mM sodium pyruvate, 4500 mg/L glucose, and 1500 mg/L sodium bicarbonate an beta-mercaptoethanol.

Are those compounds necessary?

2. Is there a protocol for neutrophil differentiation?

I found a protocol using dibutryl cAMP would this work ?

I'm working with Herpesvirus and Adenovirus. For Herpesvirus, I use Vero cell line to activate and check virus titer. For Adenovirus, I use Hela cell line.

Recently, I could activate them, but can not have high titer. The titer i need around 10^9 or 10^10 TCID50. But I just have around 10^5 TCID50.

I use this protocol to activate virus:

1. Cell cultured in 75cm2 flask for 2- 4 days, until 90 - 10% confluent.

2. Wash 2x with PBS.

3. Infect 3ml of virus stock for each 75cm2 flask for 90mins, rock the flask every 15 mins to deposit all cells in flask.

4. After 90 mins incubation with 3 ml of virus stock. Add more 20 ml of medium (DMEM+2%FBS+1%P/S + 44mM Sodium bicarbonate).

5. Incubate flask for 2 - 3days until 90 - 100% CPE appears.

6. Freeze (-70 degree) and thaw (4 degree) 3x to lyse cell.

7. Cfg 1500xg, 4 degree, 10 ml, collect supernatant for virus stock.

Could you please show me specific protocol to increase these viral titer? Thank you so much.

Sodium Bicarbonate intraperitoneal dose for adult patients using PD solution containing of Lactate. How often can be used? And what are the Precautions should be taken during the administration

When I run a solution containing dissolved salts of Mg, Ca,, and Na and as well sodium bicarbonate through a Nylon based 1um filter, I noticed that pH increases.

I am concerned that some salts are removed. Do Nylon based filters do remove some ions or it could be only that H+ ions stick to te Nylon membrane?

I want to culture Spirulina in a large scale. but for example a 10,000-L medium needs to 160Kg Sodium Bicarbonate according to Zarrouk!!! I have a protocol that requires 8 g/L of Sodium Bicarbonate, which is still too much. i need a protocol to reduce the volume and cost. would you please help me?

I want to culture Spirulina in a large scale. but for example a 10,000-L medium needs to 160Kg Sodium Bicarbonate according to Zarrouk!!!

I have a protocol that requires 8 g/L of Sodium Bicarbonate, which is still too much.

i need a protocol to reduce the volume and cost. would you please help me?

Method and solvent

I'm formulating a natural deodorant and found that the 5% concentration of NaHCO3, it works great for inhibiting the scent of sweat, but about 15% of subjects get a rash with chronic usage, ie daily for weeks.

Would 1% still have an anti-microbial effect to prevent odor?

Hi, I am trying to dissolve pure reagent grade Quercetin into cell culture medium (DMEM+sodium bicarbonate+antibiotics) but it clearly does not form a solution. I've found that DMSO might serve but high DMSO concentrations are toxic for cells. Does anyone have found a non-cytotoxic method for dissolving quercetin? Thanks in advance.

I am going to be growing NK-92 Cells with the ATCC recommended media recipe below. I am trying to make frozen aliquots of all of the components for ease of making future bottles of media and the folic acid recommends dissolving in 1M NaOH. My other option I have seen is dissolving in DMSO, which I am not necessarily comfortable with growing cells in DMSO containing media long-term. Someone recommended on another post to just dissolve it straight into the media, however measuring out 4.4mg for 500 mL of media allows for much more room for weighing error than I would like.

Will using 1 M NaOH be fine for the cells provided I make the folic acid as concentrated as I can to add the least amount of NaOH possible? Or am I able to use something a little less harsh like sodium bicarbonate to dissolve the Folic Acid? Any other recommendations for cell culture help are also welcome.

NK-92 Media:

Alpha Minimum Essential Medium w/o ribonucleosides and deoxyribonucleosides

2 mM L-glutamine

1.5 g/L sodium bicarbonate

0.2 mM inositol

0.1 mM 2-mercaptoethanol

0.02 mM folic acid

100-200 U/ml recombinant IL-2

12.5% Horse Serum

12.5% Fetal Bovine Serum

Hey there,

I'm using commercially available Drabkin's reagent to measure the hemoglobin content of blood photometrically. The reagent I bought contains sodium bicarbonate, potassium ferricyanide and potassium cyanide. In my understanding, all different forms of hemoglobin (except sulfhemoglobin) are converted to methhemoglobin by potassium ferricyanide in a first step which reacts then with potassium cyanide to cyanomethemoglobin which shows a significant absorbance at 540 nm.

But sometimes I see a second peak around 575 nm (indicated by the red arrow in the graphic), sometimes very weak but sometimes also very strong and I wonder what it could be. Could this be some remainings due to uncomplete reaction or could it maybe be due to poor quality of the sample?

Thank you for your advice!

I have noticed some studies report using sodium bicarbonate and some report using sodium carbonate to determine total phenolic content of their samples. My question is does it matter which one to use and what concentration as the range is anywhere from 7.5% to 20% solution. Thank you in advance for any suggestion.

I'm working with trace amounts (say, 1 microgram) of sterols such as dinosterol, stigmasterol, and cholesterol, and have been trying to acetylate with acetic anhydride and pyradine. This is a standard procedure our lab has performed for years. Transfer the sterol to a vial insert with toluene, evaporate, add 20 ul of anhydride and 20 ul of pyridine, heat to 70C for half an hour, cool, evaporate reagents. For several months now, though, the sterol acetate yield from this step has been 20-30%, instead of our typical 90-100.

The confusing part is that it appears that sterol is being destroyed/converted, not just failing to react. When I acetylate a sterol standard and get 30% yield of the acetate, I don't see un-derivitized sterol in my chromatogram. Furthermore, if I sylilate the residue with BSTFA/TMCS, I do NOT recover the un-acetylated sterol as a TMS ether, either. If I sylilate an un-reacted sterol standard, however, I get 100% of what I expect as the TMS ether (so I know it's not a problem with my standard being more dilute than it should be). This does NOT happen with n-alkyl alcohols. nC21-OH, for example, acetylates with 100% yield just fine.

I have tried new containers of acetic anhydride and pyridine. I have tried diluting the mix with toluene and/or changing the ratios of anhydride/base. I have tried changing glassware, or using teflon. I have tried purging the reaction vial with nitrogen and argon. I have tried substituting triethylamine for the pyridine. I have tried water/DCM phase separations followed by drying of Na2SO4 as a workup instead of simply evaporating the reaction mixture. I have tried pre-deactivating my glassware. I have tried letting the reaction proceed at room temperature overnight. I have tried leaving the reaction at 70C overnight. I have tried a different method that uses toluene, acetic anhydride, and sodium bicarbonate as a sparingly-soluble base that keeps the byproduct acetic acid quenched. I've compared results on a GC-FID with a PTV inlet, a GC-MS with a split/splitless, and someone elses GC-FID with split/splitless in another lab.

Nothing has worked.

Does anyone, in the name of all that is good, have any idea where my sterol might be going? Or how to proceed to fix this? Simple anhydride:pyridine at 1:1 was always effective in the past, and the lack of un-reacted sterol tells me it's not a reaction rate problem that a catalyst will solve.

Regardless, I currently have test reactions running that include DMAP and n-methylimidazole as catalysts. I haven't worked them up yet, but for former turned deep yellow and the latter turned deep red. The reactions in which I used TEA as the base also turned yellow, and left behind a large amount of oily residue. Are these warning signs or telling indicators of some sort of oxidative problem?

In a couple of days I will have materials for methods using silver triflate or montmorillanite clay as catalysts, but I have no reason to suspect these will go any differently.

Edit: Edited to indicate that, in contrast, straight chain saturated alcohols DON'T seem to have any problem acetylating with 100% yield under normal conditions.

I would like to know what else need to be added to make complete medium that can be used in 5% CO2 incubator, do we have to add Non Essential Animo Acids and Sodium Pyruvate, and HEPES apart from sodium bicarbonate? and how much amount Sodium bicarbonate needs to be added to prepare 1L medium to be used in 5% CO2 incubator. To be more specific, Gibco Minimum Essential Medium powder, catalog number 61100-061

I’m having trouble in culturing THP-1 for over 2 months now. I followed the protocol you provided in your web page. We are establishing this cell line in our lab so we got a frozen vial from different research center located in different city and the vial was transferred in dry ice. When I got the vial it says that viability is 99%. And I couldn’t get any information about how that vial was cryopreserved since the center are not working in this cell line anymore and the vial was stored at Liquid nitrogen since 2005. Mean wile I thawed the vial using RPMI Medium 1640 from Gibco with (+ 4.5 g/L D-Glucose + 2.383 g/L HEPES Buffer + L-Glutamine + 1.5 g/L Sodium Bicarbonate + 110 mg/L Sodium Pyruvate) and followed the steps below:

Day 1(29% viability):

1. I prepared complete media + 20% FCS + Glutamine in case was degraded in our media.

2. I thawed the vial for 2 minutes

3. I decided to leave it settle down for 2.5 hours in the incubator

4. I centrifuge for 5 minutes at 300g

5. I discard supernatant and resuspend pellet in 5 ml of media

6. Transfer cell content into new 25 ml flask and keep upright in the incubator and see after 48 hours

7. From beginning I noticed that the live cells number is very small compared to dead ones.

8. Viability was 29%. HOWEVER, Viability of the vial was 99% and method and condition of cryopreserving of that vial was unknown.

Day 2(27% viability) 3days (from thawing):

1. I prepared complete media + Glutamine

2. I decided not to do any centrifugation , just add new 10 ml complete media+ Glutamine

3. Leave flask in incubator over weekend

Day3 (6% viability) 6days (from thawing):

1. I notice that number of live cells has dropped and viability is 6%

2. I notice that media is turbid and very small thing appear under microscope ( may be Dead Cells Debris)

3. I centrifuge for 5 minutes at 300g

4. I discard supernatant and resuspend pellet in 5 ml of media ( 2 ml of old medium + 3ml of new medium)

5. Transfer cell content into new 25 ml flask and keep upright in the incubator and see after 24 hours

Today … 7 days (from thawing):

1. I centrifuge for 5 minutes at 300g

2. I discard supernatant and resuspend pellet in 5 ml of new media (no old media)

3. Transfer cell content into 6 well culture plate and keep in the incubator and I will see after 24 hours

I would appreciate the chance to have your advice in my steps and how my culture of Cell look like.

I have attached my cell culture for all 4 days.

On two visits to Tenerife my long term digestive problems stop within a day. On investigating, I find that adding Silicon 30mg /litre and sodium bicarbonate to bring the total alkalinity to 240 to match the water in Tenerife has the same effect. Why ?

It was observed that the drug released from microspheres decreased as the concentration of sodium bicarbonate increased

1-During anaerobic treatment of wastewater, sodium bicarbonate is added for alkalinity and pH control. I already know how to calculate this sodium bicarbonate dose.

2-If a energy mass balance is done around this anaerobic treatment, energy is produced in form of methane, but how could I calculate the energy entering the system in the form of sodium bicarbonate?

I used Gibco RPMI 1640 ( 4.5g/L D-Glucose, 2.383 g/L Hepes Buffer, L-Glutamine, 1.5g/L Sodium Bicarbonate, 110mg/L Sodium Pyruvate) medium supplemented with 2% Human serum for monocytes plastic adherence and then i used for macrophages culture the same medium composition plus 50ng/ml GM-CSF to generate M1 macrophages and 50 ng/ml M-CSFfor M2 macrophages.

The problem i have , is that i don't get enough macrophages after 5 or 7 days differenciation and the M1 macrophages are CD206 positve and low CD 86 positive whereas the M2 macrophages are CD206 positive and CD86 positve.

If someone has a good Media composition please share with me

After some consulting I decided to use a commercial pcr cleanup spin column kit to concentrate DNA from my CHIP for sequencing. The DNA was in elutate of sodium bicarbonate and 1% SDS. Instructions required me to dilute 6x in binding buffer, so this meant 9mL per sample column. When combined, precipitate came out, I assume this was the SDS. All of the protocols I've seen that use sodium biocarbonate and SDS to elute out dna jumped straight to purification using p/c or kits so I just moved on.

I ran the liquid through the column and near the end when I was washing them I noticed that the membrane of the tubes looked a bit crumpled and the wash buffer was dripping through it somewhat faster than in control columns and whitish precipitate was in the wash. I tried looking at the membrane at every angle but I didn't see any hole. After I eluted I noticed a tiny pellet of whitish stuff at the bottom of the tube. I'm not sure whether this was SDS or silica or what. I had the elutant tested through qubit and the samples showed very low to below threshold amounts of DNA. But this was CHIP pulldowns so this is not necessarily indicative that something is wrong. I was wondering what sort of steps I should take from here. Should just move on and see if I can create a library and do a PCR check or assume the columns were broken and try to reprocess the binding buffer waste which I kept? Multiple companies I contacted seemed to think that the spin columns should be able to withstand that volume although one of them (Thermofisher) apparently has columns with membranes that look thicker than the kit I was using.

Can we use DMEM medium for hybridoma cell culture adding Sodium bicarbonate without keep in a CO2 incubator? Because at the moment we are using MEM medium without CO2 incubator for cancer cells. Therefore, I need to clarify whether we can use DMEM in same CO2 free incubator for hybridoma cells.

I am developing whey protein beverage that sterile by retort. I have to adjust pH of product from 6.5 to above 7 to prevent protein coagulation during retort process.

I think I will use sodium bicarbonate to adjust pH. Could you give me suggestion? If you have another idea please let me know.

Thank you for your attention

Regards,

We are using above mixture for our cancer cell culture. I need to know whether we can use the same medium for hybridomas, instead of RPMI or DMEM which is mentioned in the product details. pls reply me.

I'm running a conjugation reaction to label 20nt oligonucleotides with an NHS ester fluorophore. The oligos have a 3' amino group to anchor the ester.

I dissolved the NHS ester in 100% DMSO and the oligos in 0.1M sodium bicarbonate (pH 8.4), and added them together into a single tube for overnight reacting at 4˚C.

As soon as I added the NHS ester to the oligos, I noticed a bunch of small solid particles precipitate. Is this the DNA? Will this likely result in the reaction not proceeding or getting very little conjugated oligos? Any recommendations on dissolving this precipitate?

As we know, the chemical pesticides or fungicides in the fruit are not eliminated by washing. Since these toxins are soluble in water, does the immersion of fruits in water (from 20 min for grapes to 8 h for citrus) or sodium bicarbonate (baking soda) 1%, eliminates these toxins from the fruit?

Hello everyone,

i work on various cell lines, recently i procured THP-1 cell line, which i am unable to culture in regular DMEM media with high glucose without sodium pyruvate and low sodium bicarbonate along with 50um of 2-mercapthoethanol, can anyone suggest what will be suitable culturing media for THP-1 cell line and any other components to be added for their culture.

Thank you.

I plate MDCK cells in 12-well plate and make sure next day wells are a confluent monolayer. Then I aspirate the cell culture supernatant and wash with PBS twice, and add 200 ul of tenfold virus dilutions to each well and incubate for an hour in 37 at RT for 1h and rock the plate side to side every 10 min. Then I mix and add the agar overlay which consists of sterile water, 2x MEM, 1% DEAE dextran, 5% sodium bicarbonate, 1 ug/ml trypsin-TPCK, and 2% Oxoid agar. The agar solidifies at room temperature for 15 minutes before placing in the incubator. After 24 hours, the cells are all dead. So I have tried a lot conditions (to make sure the agar was not too hot and to change ingredients of agar overlay). Finally, I found all MDCK cells without trypsin-TPCK in agar overlay survived. Does someone can tell me what happened to my trypsin-TPCK (they were prepared in 2016 and stored in -20). And Now I have ordered new trypsin-TPCK, the instruction showed it should be dissolved in 1mM HCl (ph=3), while someone said dissolved in DMEM or sterile water, I'm not sure about it. any suggestion?

Does anyone has prepared RNC(Rany Nickel catalyst) of 85-90% Purity?

We using leaching method with NiAl alloy treated with Sodium Bicarbonate and it gives only 15% purity.

What promoters,dopping should enhance the ratio?

Kindly share your thoughts, experience ASAP.

Thanks in advance.

Is it necessary to boil dialysis tubes in EDTA/sodium bicarbonate before using? or is it enough to boil it in distilled water? What is the recommended protocol for preparing dialysis tubing?

Could anyone help me to explain why my mouse embryonic cells will begin die when I add some reagents into the cell medium to differentiate ESs? Mouse ES grow on MEF. Cells will be exposed in MCDB 131 medium supplemented with 1.5 g/l sodium bicarbonate, 1×Glutamax, 10 mM final glucose concentration, 0.5% BSA, 100 ng/ ml GDF8), and 3 mM of CHIR-99021 on day 1, day 2 and days almost the same reagents. I will add some new reagents like 0.25 mM ascorbic acid and 50 ng/ml of FGF7 on day 4 and day 5. Day 6- and 7 need to add new reagents like 0.25 mM SANT-1, 1 mM retinoic acid, 100 nM LDN193189, 1:200 ITS-X , and 200 nM TPB. However, my cell will always die from day 1. Sometimes my all ES cells die after day7. Why, please help me and give me some advice.

What is the mechanism that reverses this condition ?

I am preparing a modified lectin based immuno assay. I am oxidizing my monoclonal anti-PSA antibodies (Abs) to avoid non-specific lectin attachment to the carbohydrated present of the Abs. I am following the following approach for oxidation.

1) Removal of sodium azide from Abs using desalting (spin filteration)

2) Reconstitution of Abs in PBS buffer.

3) Addition of sodium periodate solution for oxidation (20mM sodium iodate, 100mM sodium acetate, 150mM of NaCl - solution made in DI-water). Add equal amount of the solution to the Ab solution, at 4C in dark and keep it for 2 hours.

4) Removal of oxidized antibodies and reconstitution in sodium bicarbonate buffer (pH9.6). This avoids self conjugation of the Abs with themselves from the amine side.

5) Conjugation of NHS-PEG-PC-biotin linker using the generic biotinylation protocol.

6) Removal of excess biotin and blocking of excess aldehyde sites with 1M ethanolamine solution.

7) Removal of excess solution and extracting biotinylated Abs.

I want to block the oxidized Abs and want to know whether my approach will be fine ? I want to conjugate the biotin linker from the amine side using the NHS chemistry. I have seen oxidized Ab blocking with MPBH + Cys-Gly dipeptides. I am not sure whether this blocking mechanism will affect my biotinylation ?

Please advise.

While managing Patient with hyperkalemia ,we administer sodium bicarbonate !how does it work , or mechanism in action ?

For alkaline soils, Olsen's method using 1M sodium bicarbonate is recommended for determination of soil available P.

However my ICP-OES technician says that because there is Na concentration in extract, P cannot be determined.

Known solutions:

1) Use UV-visible spectrophotometer for P estimation from the extract.

2) Use Colwell P method.

3) remove Na in the soil extracts and then determine P in the extracts.

Requesting experts to suggest the best solution. I have access to ICP at convenience. UV-visible spectrophotometer is not available currently.

Kindly help me in this matter.

Merry Christmas to all!

Thanking you,

Best Regards,

Rohan

Hi, I am in need of a good growth medium for HepG2 cells. i know there are a lot of people using different types with varying additives and glucose but does anyone have experience using Williams’ Medium E liquid, sterile-filtered, suitable for cell culture, With sodium bicarbonate and L-glutamine, without phenol red ? I like to exclude the phenol red because it interferes with MTT protocols.

thanks in advance

Due to the bacterial actions, the pH of the liquid media always stay acidic (about 4 - 5). For the purpose of maintaining the pH alkaline (above 8), dilute NaOH solution was initially used to adjust the pH on daily basis, then we thought about using the sodium bicarbonate as an alternatives.

However, after adding the baking soda power in the media, the gas production  ceased, the pH stayed quit stable (about 9 above). But I am wondering if the baking soda killed the bacteria or micro-organism ?

Is there anyway we can test if the bacteria is still alive?

Does anyone have the experience of keeping the pH alkaline of the bacterial media?

Hi,

I recently bought DMEM/f12 without glutamine and sodium bicarbonate from Corning cellgro. Before using media I add glutamine, gentamicine and B27. This time media color change to orange-yellow. I checked PH and it is around 7.5. I am just wondering what is the reason of color change, is there any reaction between media and these supplements? I am not sure that I can use it or not.

Companies formulate DMEM always with 3.7g/L of sodium bicarbonate. But majority of researchers use only 5% CO2 incubators, which is ideal for media with 1.5-2.2g/L of sodium bicarbonate. 3.7g/L of NaHCO3 requires 10% CO2. So, while most of us use only 5% CO2 incubators and everyone knows this, companies must have a good reason for insisting on 3.7g/L. Do you have an idea on this? Does DMEM have to contain 3.7g/L NaHCO3 for some reason?

People in my lab also use DMEM but they didn't know DMEM requires 10% CO2. I let them know this but they are not willing to change their culture conditions as all of their experiments have been done in 5% CO2 all the time. So I decided to buy DMEM with lower NaHCO3 concentration. I found one from ATCC (1.5g/L), but they only have high glucose. I found powder forms where I can add the desired amount of NaHCO3 but I don't want to spend time on preparation, pH adjustments and sterility issues. Do you have a suggestion about what to do? Should I just ignore these questions in my mind and keep doing what many researchers have been doing wrongly (i.e. using DMEM with 5% CO2)?

Thanks in advance for your replies.

Is sodium bicarbonate installation (small amount and diluted) used for artificial airway (ETT and Tracheostomy) suctioning to remove thick secretions? Clinically and theoretically, Why and why not?

Intensivists, pulmonagists, Critical Care Nurses and researchers. 

Thank you.

A 3.5% saline solution prepared with water and sodium has a pH of 6.38.

I want to make a solution of 3.5% salinity with a pH of 8.1 how do i do this using only sodium, and sodium bicarbonate?

I need to adjust for the sodium of the sodium bicarbonate in regards to the salinity whilst maintining a ph of 8.1 and salinity of 3.5%.

The aim of this is to make a solution that will resemble sea water pH and salinity.

Hi! This protocol has been used in my lab to handle healthy rat liver samples (see below). But now, i'm going to start a project for quantifying membrane proteins in healthy and diseased human liver tissue and this doesn't feel like a safe way of doing it.

Has someone homogenized Human liver before that could share a few tips and maybe the protocol they followed?

thank you!!!

Isolation of membrane protein

Rat liver and kidney weighing were homogenized using glass dounce homogenizers in Sodium Bicarbonate with protease inhibitors. After brief centrifugation, the clear supernatant was incubated with Sodium Carbonate at 4 oC for 15 minutes. Samples were centrifuged at 100,000g for 1 hour at 4 oC. The pellet was resuspended in1xPBS containing protease inhibitors and disrupted briefly by ultrasonication. The protein was quantitated by Pierce BCA Protein Assay kit. 

Addition of sodium bicarbonate to the medium alters the pH of the broth and by which the growth and production of microbe gets altered; but , does it really alter the structural composition of the PHA synthesized?

For herbal antacid/effervescent , will sajjikshar and lemon juice powder will give the same amount of fizz as given by lab grade citric acid and sodium bicarbonate?