Cells Detachment

Easy, Gentle and Complete

I often get asked if trypsin is the best reagent to detach primary and stem cells. The problem with a trypsin is that it could lyse a percentage of the cells.

We recommend our Detachin^TM. It offers the following benefits:

• Gentle and rapid detachment.
• Maximum cell viability.
• Effective on a wide range of cells.
• No mammalian or bacterial byproducts.
• Stable at 4°C for 2 months.
• No need to wash detached cells.

Here's a recent publication referencing the use of Detachin: Yu Hou, Qi Feng, Miao Xu, Guo-sheng Li, Xue-na Liu, Zi Sheng, Hai Zhou, Ji Ma, Yu Wei,1 Yuan-xin Sun,Ying-yi Yu, Ji-hua Qiu, Lin-lin Shao, Xin-guang Liu, Ming Hou, and Jun Peng. High-dose dexamethasone corrects impaired myeloid-derived suppressor cell function via Ets1 in immune thrombocytopenia. Blood 2016 :blood-2015-10-674531; doi:10.1182/blood-2015-10-674531 ...Both PBMCs and splenocytes were cultured in complete medium for 7 days. Each culture was supplemented with recombinant human IL-6 (10 ng/mL; R&D Systems, Minneapolis, MN) and granulocyte macrophage-colony-stimulating factor (10 ng/mL; R&D Systems) in the presence or absence of DXM and incubatedin humidified air with 5%CO2 at 37°C. Cells cultured in medium alone were runin parallel as controls. Adherent cells were removed using a nonprotease cell detachment solution Detachin (Neuromics, Edina, MN). CD331 cells were isolated by anti-CD33 magnetic microbeads and LS column separation (Miltenyi Biotec, Bergisch Gladbach, Germany), per manufacturer's instructions. The purity of the isolated cells was found to be higher than 90% by flow cytometry

Figure 1. The number of MDSCs and their expression of Arg-1/iNOS in the peripheral blood. (A) The representative scatter-gram of CD11b1CD331HLA-DRlow cells within the gate of PBMCs. Histograms of Arg-1 and iNOS in CD11b1CD331HLA-DRlow cells from healthy control patients and patients with ITP before any treatment was initiated. (B) The percentage of CD11b1CD331HLA-DRlow cells in hemolyzed whole blood from primary active patients with ITP (n 5 21) and healthy control patients (n 5 18). (C-D) The expression (mean fluorescence intensity) of Arg-1 (C) and iNOS (D) in circulating MDSCs compared between patients with ITP (n 5 21) and control patients (n 5 18). Significance between the 2 groups was determined by Student-Newman-Keuls test.

We are working hard to find new ways for you to optimize your cell based assays.

Cells Detachment

Easy, Gentle and Complete

I often get asked if trypsin is the best reagent to detach primary and stem cells. The problem with a trypsin is that it could lyse a percentage of the cells.

We recommend our Detachin^TM. It offers the following benefits:

• Gentle and rapid detachment.
• Maximum cell viability.
• Effective on a wide range of cells.
• No mammalian or bacterial byproducts.
• Stable at 4°C for 2 months.
• No need to wash detached cells.

Here's a recent publication referencing the use of Detachin: Yu Hou, Qi Feng, Miao Xu, Guo-sheng Li, Xue-na Liu, Zi Sheng, Hai Zhou, Ji Ma, Yu Wei,1 Yuan-xin Sun,Ying-yi Yu, Ji-hua Qiu, Lin-lin Shao, Xin-guang Liu, Ming Hou, and Jun Peng. High-dose dexamethasone corrects impaired myeloid-derived suppressor cell function via Ets1 in immune thrombocytopenia. Blood 2016 :blood-2015-10-674531; doi:10.1182/blood-2015-10-674531 ...Both PBMCs and splenocytes were cultured in complete medium for 7 days. Each culture was supplemented with recombinant human IL-6 (10 ng/mL; R&D Systems, Minneapolis, MN) and granulocyte macrophage-colony-stimulating factor (10 ng/mL; R&D Systems) in the presence or absence of DXM and incubatedin humidified air with 5%CO2 at 37°C. Cells cultured in medium alone were runin parallel as controls. Adherent cells were removed using a nonprotease cell detachment solution Detachin (Neuromics, Edina, MN). CD331 cells were isolated by anti-CD33 magnetic microbeads and LS column separation (Miltenyi Biotec, Bergisch Gladbach, Germany), per manufacturer's instructions. The purity of the isolated cells was found to be higher than 90% by flow cytometry

Figure 1. The number of MDSCs and their expression of Arg-1/iNOS in the peripheral blood. (A) The representative scatter-gram of CD11b1CD331HLA-DRlow cells within the gate of PBMCs. Histograms of Arg-1 and iNOS in CD11b1CD331HLA-DRlow cells from healthy control patients and patients with ITP before any treatment was initiated. (B) The percentage of CD11b1CD331HLA-DRlow cells in hemolyzed whole blood from primary active patients with ITP (n 5 21) and healthy control patients (n 5 18). (C-D) The expression (mean fluorescence intensity) of Arg-1 (C) and iNOS (D) in circulating MDSCs compared between patients with ITP (n 5 21) and control patients (n 5 18). Significance between the 2 groups was determined by Student-Newman-Keuls test.

We are working hard to find new ways for you to optimize your cell based assays.

Everything You Wanted to Know About Stem Cells

Unforgettable way to learn about Stem Cells.

Rap on Dr. Jonathan Garlick Rap on.

If you are doing stem cell related research, check out our:

hNP1™ Human Neural Progenitor & hN2™ Neuron Discovery Kits Derived from H9 (WA09) ECSs-Consistent, Easy to Use & Cost Effective	Neural Stem Cells Media
hMPro™ Human Mesenchymal Progenitors (hMPCs) hESC Derived	Human Mesenchymal Stem Cells (hMSCs) hMSCs Derived from Umbilical Cord Blood
MSCGro™ Mesenchymal Stem Cell Media Proven and Potent Culture Growth and Differentiation Media for Mesenchymal Stem Cells (MSCs)	Cell Cryopreservation Media Maximizes cell recovery, attachment and growth
Expansion/Differentiation Kits SC to Oligodendrocytes and Dopaminergic Neurons Kits	Bioluminomics™ In Vitro Assays Brought to you by HemoGenix®
3-D Cell Based Assay Solutions Nanofibers, Hydrogels and Extracellular Matrix (ECM) Proteins	Stem Cell Research Antibodies
Stem Cell Research Proteins	Rat Neurosphere Neuroprogenitor Tissue Tissue provided live, unseparated, fresh from E18 rat cortex/hippocampus including subventricular zone.

Shiny, New GF nanoparticle labeled hMSCs

Human Mesenchymal Stem Cells-Green Fluorescent Nanoparticle-labeled-These cells are generated by chemical transfection of CdSe/ZnS nanoparticles, 100 nm in diameter and show a strong fluorescent signal (Excitation maximum= 515 nm; Emission maximum= 520-550 nm) detectable by standard FITC filters.

Image: FITC analysis of cells during fourth passage following transfection. Image obtaining using FITC filter on Olympus CKX41 microscope at 200 X equipped with a Retiga 2000 digital camera and Q-Capture software program. Scale bar is 25 micrometers. Fluorescent nanoparticles are less prevalent within the cytoplasm than earlier passages. Technical Information & Data.

They retain GFNP labeling through 3+ passages and can be differentiation into chondrogenic, adipogenic or osteogenic lineages. Excellent for regeneration, toxicity and related studies. Technical Information & Data.

Image: Osteogenic differentiation of GF nanoparticle-labeled MSCs. Image obtaining using phase contrast Olympus CKX41 microscope at 100 X equipped with a Retiga 2000 digital camera and Q-Capture Pro software program. Scale bar is 25 micrometers. Note the presence of mineral deposits indicating differentiation into osteoblasts. Chondrogenic and adipogenic differentiation was also apparent.

• All Stem Cell Research Reagents.

True Physiologic Conditions in Cell Culturing

The Petaka Advantage!

A cornerstone of Neuromics' strategy is to enable better science by finding ways to help our customers and collaborators to both improve and lower the overall costs of their cell based assays.

We embrace new technologies if they prove capable of providing cell culture environment that more closely mimic in vivo environments. This enables basic and drug discovery researchers learn more about the true potential of targets. More informed decisions early in the process reduce downstream costs.

This is why the use of engineered mini bioreactors with Ducted Respiratory Chambers (DRCs) like the Petaka® G3 LOT needs to be considered. (detailed information @ Ducted Respiratory Chamber Bioreactors.© 2012 Genetic Engineering & Biotechnology News All Rights Reserved). Instead of attempting to impose “normal” gas conditions on the cells through active incubator controls, this system passively allows cells to maintain their own oxygen, carbon dioxide, and humidity levels.

Image: Petaka's DRC design. The cell culture chamber is isolated on the injection side from the atmosphere by a self-sealing silicone injection port that allows the closed introduction of media and cells, including most eukaryotic cells types, small early-stage embryos, tissue fragments, and even needle biopsies.

Because the cells control their own gas environment, there is little effect from outside gas conditions, and the DRC is effective in atmospheres from 500 meters below sea level up to elevations of 4,000 meters, and in relative humidity levels between 10% and 100%.

Figure: Self-regulating gas management in the DRC. Cells consume O₂, with restricted O₂ ingress, causing a first proliferative cell phase to evolve into a second differentiating cell phase for protein production and gene expression.

The DRC changes the paradigm of cell culture, replacing nearly a century of active attempts to humanly intervene to manipulate gas exchange with a self-regulating design driven by natural laws of diffusion to create dependable, and truly physiologic, gas environments.

Laryngotracheal Transplantations Using Nanofiber Solutions Scaffolds

Nanofibers Solutions work in transplants-imagine how well they will work in your 3-D based cell based assays.

3-D Cell Based Assays for Drug Discovery are the future. Like any new model, adoption rates are a function of how well the new solutions works. "The proof is in the pudding".

Here're highlights of a historic event based on transplants using nanofiber engineered laryngotrachea : Collaboration between Nanofiber Solutions and the Karolinska Institutet produces first synthetic laryngotracheal implants seeded with the patient’s stem cells to be successfully transplanted into human patients in Russia.
COLUMBUS, Ohio, June 26, 2012 – Nanofiber Solutions, LLC, an Ohio-based developer, manufacturer and marketer of 3-D synthetic scaffolds to advance basic research, tissue engineering and regenerative medicine announced today the first and second successful transplants of its tissue engineered laryngotracheal implants seeded with cells from the patients’ bone marrow.

The surgeries were performed June 19th and 21st at the Krasnodar Regional Hospital (Russia) by Dr. Paolo Macchiarini, Professor of Regenerative Surgery at the Karolinska Institutet (Stockholm, Sweden), and colleagues. Dr. Macchiarini led an international team that included Dr. Vladimir Porhanov, head of Oncological and Thoracic Surgery at Kuban State Medical University (Russia), Dr. Jed Johnson, Nanofiber Solution’s Chief Technology Officer who created the synthetic organs, Harvard Bioscience (Boston, USA) who produced the bioreactor, and Dr. Alessandra Bianco at University of Rome, Tor Vergata, who performed mechanical testing during scaffold development.

Both patients, a 33 year-old mother from St. Petersburg and a 28 year-old man from Rostov-on-Don, were in au to accidents and suffered from a narrowing of the laryngotracheal junction for which they already had failed previous surgeries. Transplantation was the last option for the patients to have normal quality of life. Immediately following transplantation, both patients were able to speak and breathe normally.

 Nanofiber Solutions, lead by Dr. Johnson, designed and built the nanofiber laryngotracheal scaffolds specifically to match the dimensions of each patient’s natural larynx and trachea, while Harvard Bioscience provided a bioreactor used to seed the scaffold with the patients’ own stem cells.  Although this procedure represents the world’s first and second successful use of synthetic synthetic laryngotracheal implants, it is Nanofiber Solution’s second and third successful organ implants using their synthetic scaffolds within the last year.

Nanofiber Solutions’ scaffolds mimic the body’s physical structure and allow for a more successful seeding, growth and differentiation of stem cells. Because the cells used to regenerate the larynx and trachea were the patients’ own, doctors report there has been no rejection of the transplants and the patients are not taking immunosuppressive drugs. (more).

Capabilities of 3-D nanofiber scaffolds for cell based assays:

Human brain tumor biopsy showing migrating tumor cells along the alligned nanofiber.

Nanofibers are optically transparent to allow for live-cell imaging and real time quantification of cell mobility using an inverted microscope

Nanofibers mimic the 3D topography found in vivo which produces a more realistic cellular response to therapeutics.

More realistic cellular behavior means you can use fewer animals and decrease time-to-market for drug discovery and development.

Nanofibers can easily be coated with ECM proteins using existing protocols for standard lab ware.

Cells can be easily removed for protein or gene analysis using trypsin, EDTA, etc.

We will continue posting relevant press releases, pubs and data that prove the capabilities of these important solutions.