NutriStem® V9 XF FAQ's

Does NutriStem® V9 XF contain any serum or purified/recombinant serum proteins?

NutriStem® V9 XF contains clinical grade human serum albumin and recombinant proteins.

What is the difference between NutriStem® hPSC XF and NutriStem® V9 XF?

NutriStem® hPSC XF is optimized for Matrigel and Laminin521 matrix. The NutriStem® V9 XF is optimized for Vitronectin matrix.

Is the product configuration of NutriStem® V9 XF different than NutriStem® hPSC XF?

Yes. NutriStem® hPSC XF is a complete ready to use medium. NutriStem® V9 XF is composed of frozen basal medium and a frozen 500X growth factor supplement.

Does NutriStem® V9 XF allows weekend free feeding regime?

Yes. Best results observed when cells are split twice weekly. To enable a weekend-free culture schedule, cells would be split on Monday and Thursday with full media exchanges on Wednesday (48h post seeding) and Thursday, followed by a passage on Friday. In this example, the Friday passage should be done into twice the standard volume with a medium change 48h later.

What is the timing for medium changes and passaging?

Optimal results are observed when the cells are split twice weekly. At the first day after cells seeding it is best to not move the plate or change medium. Approximately 2 days after plating change the medium daily with 3ml/well complete NutriStem® V9 XF medium until the colonies are large enough to passage.

What is the recommended matrix for NutriStem® V9 XF?

Best results obtained with Vitronect. NutriStem® V9 XF may also be used with Basement Membrane Matrix (Matrigel).

What is the recommend vitronectin for this culture system?

All major vitronectin suppliers: Peprotech, SCT (Primorigen), Life (advanced biomatrix) and Millipore have been used successfully with NutriStem® V9 XF. The precoating- free application works ONLY with Vitronectin ACF.

What is the recommended passage method for NutriStem® V9 XF?

We recommend passaging the hPSC as small aggregates using 0.5mM EDTA solution. Dilute 0.5M EDTA (01-1862-1) 1:1000 in calcium and magnesium free DPBS. We do not recommend single cell dissociation.

Do I need to add a ROCK inhibitor during passaging?

Addition of ROCK inhibitor is not required for routine work with NutriStem® V9 XF.

What are the recommended passaging ratios?

Passaging ratios depend on the cell line you are using. Usually a splitting ratio of 1:8-1:20 every 4 days is required. Lower split ratio (1:3-1:6) may be needed during recovery from passage or during adaptation.

How shell I thaw NutriStem® V9 XF of complete medium?

Thaw NutriStem® V9 XF basal medium at room temperature or at 2–8°C. Thaw NutriStem® V9 XF supplement mix on ice. Do not thaw either basal medium or supplement mix in a 37°C water bath.

What do I need to do to prepare NutriStem® V9 XF complete medium?

Thaw NutriStem® V9 XF basal at 2–8°C, and supplement mix on ice. Add supplement mix (1ml) to 500ml basal medium and mix gently. Complete medium may be used freshly, up to 2 weeks if stored at 2-8ºC or aliquoted for later use. freeze aliquots of complete NutriStem® V9 XF and use within 6 months.

Can I thaw NutriStem® V9 XF basal medium at 37°C water bath?

No, do not thaw the NutriStem® V9 XF basal in a 37°C water bath. Best results are observed for thawing at room temperature. You may also thaw at 2–8°C.

What is the shelf life of NutriStem® V9 XF after supplementation (Complete medium)?

Best results are observed when complete NutriStem® V9 XF is used within 2 weeks from preparation.

Can I warm complete NutriStem® V9 XF in a 37°C water bath for daily use?

No, do not warm NutriStem® V9 XF at 37°C. Best results are achieved when the medium is allowed to slowly reach room temperature or equilibrated at 37°C CO2 incubator in cultureware prior to cell seeding.

Can I use NutriStem® V9 XF in my reprogramming and gene editing workflows?

The compatibility of NutriStem® V9 XF with reprogramming or gene editing workflows has not been tested.

Can I cryopreserve cells cultured in NutriStem® V9 XF?

Cells cultured routinely in NutriStem® V9 XF can be cryopreserved and revived efficiently.

Does NutriStem® V9 XF supports long-term cultures?

We successfully maintained hPSC for up to 30 passages in multiple cell lines. Pluripotency marker expression (Tra-1-60, SSEA4, Sox2, Oct-4, and Nanog), differentiation potential, and normal karyotypes are all maintained in long-term culture with of NutriStem® V9 XF.

Does routine culture in NutriStem® V9 XF have any effect on the karyotype of my cells?

We have observed normal karyotypes in multiple PSC lines during long-term culture up to 30 passages.

Can I use PSCs that are long-term cultured in of NutriStem® V9 XF for downstream differentiation?

We have seen no effect on downstream differentiation potential for cells cultured in NutriStem® V9 XF for up to 20 passages. Tri-lineage potential has been demonstrated from embryoid bodies.

Have you confirmed tri-lineage differentiation potential from PSCs cultured in NutriStem® V9 XF?

The potential for cells from NutriStem® V9 XF cultures to differentiate into cells of ectoderm, mesoderm, and endoderm lineages is unaffected by long-term culture in NutriStem® V9 XF. This has been confirmed using spontaneous differentiation (embryoid bodies formation) performed in basal NutriStem® V9 XF.

Will I see a morphological difference between cells cultured in NutriStem® V9 XF and cells cultured in other medium?

Cells cultured in NutriStem® V9 XF maintain the expected PSC morphology, with compact, homogeneous colonies, defined edges, and a high nucleus-to-cytoplasm ratio.

Do you see any change in pluripotency marker expression from PSCs cultured in NutriStem® V9 XF?

We have observed no significant change in pluripotency marker expression in multiple cell lines over 30 passages in NutriStem® V9 XF.

Is it necessary to pre-coate cultureware?

No. Precoating free procedure is optional. In this procedure, Vitronectin ACF added directly to NutriStem® V9 XF culture medium before cells seeding.

Will I see a morphological difference between cells cultured in the precoating free procedure and NutriStem® V9 XF medium vs. the cells cultured in the traditional NutriStem® V9 XF and Vitronectin coated wells?

Cells cultured in the precoating-free preocedure using Vitronectin ACF and NutriStem® V9 XF maintain the expected PSC morphology, with compact, homogeneous colonies, defined edges, and a high nucleus-to-cytoplasm ratio. We observed nearly similar expansion rate over 30 passages. in addition, pluripotency marker expression (Tra-1-60, SSEA4, Sox2, Oct-4, and Nanog), differentiation potential, and normal karyotypes are all maintained in long-term culture using the precoating free procedure and NutriStem® V9 XF.