Picture Blog: A Short Path from Human mRNA-iPSCs to Neurons in Record Speed

Traditional differentiation protocols use embryoid body (EB) formation as the first step of lineage restriction to mimic early human embryogenesis, which is then followed by manual selection of neuroepithelial precursors. This procedure is tedious and often inconsistent. We have developed a novel neural differentiation scheme that directs human iPSCs (created with the Allele 6F mRNA reprogramming kit) that progressed, as attached culture, to neural precursor cells (NPCs) in just 4-6 days, half the time it typically takes by other methods. From NPCs it takes about another 5-6 days for neural rosettes to form (see figures below); upon passage, cells in neural rosettes differentiate into neurons in 24 hours.

The neural progenitors at the rosettes stage can be stocked and expanded, before differentiated into different types of neurons. We are working on specifically and efficiently different these neural progenitor cells into dopaminergic, glutamatergic, GABAergic, and other types of sub-types of neurons with Allele’s technologies (Questions? email the Allele Stem Cell Group at iPSatAllelebiotech.com).

Neural rosettes formed efficiently in wells without going through EB.

neural rosettes formed as attached cells in less than 2 weeks

Human iPSC-derived neurons are created in a short regimen developed at Allele Biotech

Neurons appear from precursor cells shortly after the rosette stage

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Wednesday, February 5th, 2014 iPSCs and other stem cells, Open Forum No Comments

Allele Biotech Acquires BioCarta’s Distribution Business

Allele Biotechnology & Pharmaceuticals, Inc. is pleased to announce that as of January 1st, 2014, it has acquired the distribution business of BioCarta. This transaction will strengthen Allele Biotech’s presence in the antibody field, enhancing a broad customer and partnership base to further its plan in clinical diagnostics. Biocarta has been a leader in the field of gene expression for 10 years and has contributed immensely through its world leading effort of charting molecular biology pathways. The gene function maps published by Biocarta have been used and referred to by the NIH through the National Center for Biological Information and National Library of Medicine.

Among its well-regarded distribution business, for the past 11 years BioCarta had been the US and Canada’s exclusive distributor of Immune Function Assay Kits for Flow Cytometric Analysis by Glycotope Biotechnology, GmbH. These kits are clinically approved blood cell diagnostic assay products that are also heavily used for non-clinical blood studies.

Allele Biotech has recently launched a major business plan to create a large number of nano antibodies (nAbsTM). The nAbTM line will be great research tools for immunoprecipitation (co-IP), immunohistochemistry (IHC), Western blotting, co-crystallization, biologics purification. Additionally, nAbsTM willbe suitable for diagnostic assays because the single domain antibodies derived from camelid family animals are sturdy, specific, and low-cost. The inclusion of the BioCarta distribution channels in the antibody and pathway reagent fields will help speed up Allele Biotech’s expansion.

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Thursday, January 16th, 2014 Allele Mail Bag No Comments

Picture Blog: Naive Human Pluripotent Stem Cells Regrown From Allele’s iPSCs

As we blogged a month ago, the Hanna lab recently published a paper in Nature describing that human ESCs or iPSCs, which typically resemble more of mouse EpiSCs (epiblast stem cells) than ground state mouse stem cells, could be converted to naïve pluripotent stem cells if grown in a stem cell medium that includes hLIF, JNKi, and p38i.  The figure here shows that the reported system did perform well when we at Allele Biotech tested growing our banked iPSCs under similar conditions.  The colonies grown in naive stem cell conditions (B) did become dome-shaped when cultured for longer period of time; when transferred back into regular stem cell medium, the once naive-looking iPSCs formed tighter and “cleaner” colonies than typical “primed” human iPSC colonies.

A, Primed human stem cells: mRNA-iPSC line J-1 grown on CellStar-coated surface and in E8 medium. The cells have human iPSC morphology of being compact in size and in "shiny" colonies. B, Naïve human stem cells: J-1 iPSCs shown 2 days after switching to a medium similar to the Naïve Human Stem cell Medium (NHSM). Compared to primed stem cells in A, the naïve stem cells are more flat and transparent, with no spontaneous differentiation on the edges.

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Thursday, December 5th, 2013 iPSCs and other stem cells No Comments

We at Allele Biotech encourage you to help those in need

Typhoon Haiyan (Yolanda) has devastated vast areas in the Philippines and the local people are still suffering, particularly those who had minimum resources even before the disaster hit. Our company’s representatives’ recent trip to the country created a special bond between our employees and the people in the Philippines, and we will do what we can to help, including directly donate to agencies like the Philippine Red Cross. We encourage our clients, colleagues, and friends to do the same. Thank you.

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Wednesday, November 20th, 2013 Allele Mail Bag No Comments

New Allele Biotech Publication on Stem Cells

Feeder-Free Reprogramming of Human Fibroblasts with Messenger RNA
Current Protocols in Stem Cell Biology • November 13, 2013
DOI: 10.1002/9780470151808.sc04a06s27

Authors: Luigi Warren, Jiwu Wang

This unit describes a feeder-free protocol for deriving induced pluripotent stem cells (iPSCs) from human fibroblasts by transfection of synthetic mRNA. The reprogramming of somatic cells requires transient expression of a set of transcription factors that collectively activate an endogenous gene regulatory network specifying the pluripotent phenotype. The necessary ectopic factor expression was first effected using retroviruses; however, as viral integration into the genome is problematic for cell therapy applications, the use of footprint-free vectors such as mRNA is increasingly preferred. Strong points of the mRNA approach include high efficiency, rapid kinetics, and obviation of a clean-up phase to purge the vector. Still, the method is relatively laborious and has, up to now, involved the use of feeder cells, which brings drawbacks including poor applicability to clinically oriented iPSC derivation. Using the methods described here, mRNA reprogramming can be performed without feeders at much-reduced labor and material costs relative to established protocols.

Allele iPSC Service and Technology Licensing Contact: http://www.allelebiotech.com/cell-line-and-culture-services/#ips-line

New Allele Product of the Month: FP-nAb™ products for 100% pull-down

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Wednesday, November 13th, 2013 iPSCs and other stem cells No Comments