Expanding the Camelid Antibody Product Line

While Chromotek GFP-Trap resin has become one of the best sellers from the Allele Biotech’ Camelid Antibody (VHH antibody) product line, more products have been added that will prove to be great tools for GFP-related research.

GFP is a powerful tool to study protein localization and dynamics in living cells. However, the photo stability and the quantum efficiency of GFP are not sufficient for Super-Resolution Microscopy (e.g. 3D-SIM or STED) of fixed samples from cells expressing GFP-fusion proteins to visualize specific structures. Furthermore, many cell biological methods such as HCl treatment for BrdU-detection, the EdU-Click-iT™ treatment or heat denaturation for FISH lead to disruption of GFP signal.

Now we offer our GFP-Trap Booster for reactivation, boosting and stabilization of GFP, suitable for acquiring strong and long lasting signals from GFP-fusion proteins. It is based on a specific GFP-binding protein as in GFP-Trap but coupled to the fluorescent dye ATTO 488 (from ATTO-TEC). For information, please read the product description of this week New Product of the Week: GFP-Trap booster, ABP-CM-GBOOSTR, http://www.allelebiotech.com/shopcart/index.php?c=221&sc=158

Promotion of the week: All mTFP1 and mWasabi fusion plasmids are 30% off for this week only

Preview of future new product: a similarly high quality product, the RFP-Trap that pulls down DsRed derived proteins including mRFP1, mCherry, mOrange, mPlum but also mRuby and RFP-tagged fusion proteins.

Tags: anti-GFP, Chromotek, GFP coIP, GFP-Trap, mCherry, mOrange, mPlum, mRFP1, mRuby RFP-tagged fusion proteins, nti-RFP, pulldown, RFP-Trap, VHH antibody

LoxP 4-in-1 iPS Factor on Lentiviral Vectors for Efficient Reprogramming

Putting 4 iPS factors on one lentiviral vector, separated by 2A peptides, has appeared to be more efficient in generating iPS cells than having all 4 factors on individual viruses, at least in a number of cases. Stem cell-like colonies start to appear in about 2 weeks using Allele Biotech’s 4-in-1 lentivirus. In addition to the concerted effects from Oct3/4, Sox2, c-Myc, Klf4, it is also believed that the coordinated silencing of these factors after reprogramming help forming iPS colonies.

The 4-in-1 lentivirus from Allele Biotech contains loxP sites that can be used to remove the 4 cDNAs if so desired. For convenience, a new product kit is offered starting this week to include lenti-nCre in a kit with the 4-in-1 iPS viral products.

New Product of the Week 04-11-10 to 04-18-10: 4-In-One-Vector: Human OSKM Lentiviral Paticles (Oct3/4, Sox2, Klf4 and c-Myc) and Cre Lentiviral Particle kits, Cat # ABP-SC-LVI4IN1C1 or ABP-SC-LVI4IN1C5

Promotion of the Week 04-11-10 to 04-18-10: Single vial 4-in-1 is offered only this week. This product has been well established and validated, one of the reasons smaller packages are not normally offered. As a matter of fact, every batch of the 4-in-1 iPS lentivirus has been sold out.

Update note: Lentivirus inserts into the host chromosome, and is gradually being replaced by footprint-free reprogramming reagents, the best being Allele Biotech’s enhanced mRNA reprogramming factors that feature a patent-pending fusion gene.

Tags: 4-in-1, c-myc, cre, iPS, iPSCs, Klf4, lentivirus, loxP, Oct3, repgramming, silencing, Sox2

NIH Announces SHIFT SBIR Grants to Help Academic Researchers Get Jobs in Biotech

The National Institutes of Health (NIH) just announced a new type of Small Business Innovative Research (SBIR) grants. Called SHIFT SBIR, these grants are designed to “(1) to foster research that is translational in nature and (2) to transform academic scientific discoveries into commercial products and services,” according to the NIH announcement, and to also facilitate licensing of intellectual properties from academic institutions as well as promote better access to academic resources. The PI transitioning from the academic institution must be primarily employed by his/her research institution at the time of application and must be primarily employed (more than 50% time) by the company by or at the time of award.

One very attractive aspect of these grants is that they mean more money than standard SBIR grants. Up to $200,000 total costs per year and time periods up to 2 years may be requested for Phase I. Well-justified budgets up to $750,000 total costs per year and time periods up to 3 years may be requested for Phase II. That is sufficient for a good researcher to build a team to do research in one direction within pretty much any small company setting.

A little background about SBIR grants: SBIR programs sponsored by federal funding agencies including the NIH, NSF, DOE, FDA, the military departments, etc. have been a major source of funding for many biotech companies like Allele Biotech during their startup phases. SBIR grants can also be used to facilitate continued research and help business expansion even as the company grows. As an example of the effects of SBIR grants, Allele Biotech obtained 5 such grants from 2000 to 2003 and built a company from just ideas to one with a patent in RNAi, an out-licensing deal with Promega, a product line in oligo synthesis, and a structure that helped launch currently ~1,500 products since 2004. We then carried out 2 more SBIR contracts for the NIH from 2007 to now, which moved us into the field of special antigen production, iPS using Bacmam systems, viral packaging services, and hopefully more advanced antibodies in the pipeline.

The link to the full NIH announcement is here.

To read more blogs on SBIR related topics, click here.

The current topics of SHIFT SBIR solicitation is listed below for Allele Blog viewers’ convenience:

• Applying opportunities in genomics and other high throughput technologies to understand fundamental biology, and to uncover the causes of specific diseases
• Translating basic science discoveries into new and better treatments
• Development of diagnostics, preventative strategies and therapeutic tools
• Development and clinical evaluation of biomarkers for alcohol exposure and alcohol-induced tissue injury
• Therapeutic development for alcoholism treatment
• Diagnostic assessment and treatment of alcohol use disorders and comorbidity
• Alcohol biosensors and data analysis systems
• Prevention, diagnosis, and treatment of fetal alcohol spectrum disorder and alcohol-related birth defects
• Minimal dose post-exposure vaccine for rabies
• Immunotherapy to kill HIV-infected cells
• Asthma therapeutic vaccine
• Novel antifibrotic therapies for progressive liver failure
• Diagnostic measurement devices or methods for assessment of urinary leakage and incontinence
• Therapeutics for diabetic wound healing
• Pediatric formulations
• Robust diagnostic biosensors for infants
• mHealth tools for assessing and addressing health in children and families
• Wearable diagnostic and therapeutic devices for physiologic monitoring and interventions
• Wearable biosensors for persons with genetic sensitivity to environmental factors
• Therapeutic interventions for persons with physical and developmental disabilities
• Advancement of novel botanical therapies for effective symptom management of non-life-threatening conditions
• Development of interactive technologies to improve and expand delivery of mind/body interventions
• Discovery of improved methodology for the characterization of plants and their secondary metabolites
• Development of standardized, objective methods to assess patient adherence to specific CAM treatment interventions;
• Development of devices/tools to assess consistency and fidelity of practitioner approaches and other aspects of protocol implementation
• Virtual settings or online tools for clinician training and implementation of fidelity monitors
• Development and validation of enhanced patient-reported outcome assessment tools for CAM (e.g. new user (clinician, researcher, and/or patient/study volunteer)–friendly interfaces, methods to improve compatibility with research and health informatics systems currently in use)
• Development of measurement tools for assessing expectancy for effects of CAM mind-body medicine, acupuncture, and manual therapy interventions
• Novel technologies that enhance/track/monitor “real time” adherence to drug abuse (and HIV+) treatment regimens
• Technology to improve the efficacy of substance abuse treatment, treatment adherence, and reduce recidivism among criminally-involved patients
• Mobile and/or internet technology based treatment interventions to augment traditional substance use disorder (SUD) treatments and their outcome
• Technologies and/or devices to boost medication adherence for SUD patients
• Technology-based treatment platforms to standardize interventions and to make them more community-friendly
• Integrate item response theory and computer adaptive testing in measures of addiction liability.
• Brief screening tools to assess relapse risks in and out drug treatment settings
• Use of the internet to link community based outreach and HIV testing services to facilitate access by drug users and their sex partners in neighborhood settings.
• Development of novel therapeutics, diagnostics, and devices for treating heart, lung, blood and sleep diseases and disorders
• New or improved measures, analytical methods, and instruments for gene expression in individuals with heart, lung, blood, and sleep disorders and diseases
• Health-care systems and outcomes research, including development of new quality measures for evidence-based heart, lung, blood, and sleep health care
• Models of behavior modification and other approaches to behavior change related to heart, lung, blood, and sleep diseases and disorders
• Devices and technologies to prevent cardiac ischemia/reperfusion injury
• Vaccines for the prevention or treatment of heart, lung, and blood diseases
• Non-invasive methods to diagnose DVT and PE
• Technologies and strategies to advance cellular therapies for heart, lung and non-malignant blood diseases
• Therapies to treat hematologic diseases and cytopenic states
• Technologies for in vitro reduction, inactivation or removal of microorganisms and other infectious moieties from blood, blood components, and plasma derivatives
• Development of products, technologies and services to diagnose, treat and/or prevent skin and rheumatic diseases, muscle disorders, and joint and bone diseases

Tags: biotech industry jobs, career in biotech, NIH grants, SBIR, start up, translational research

Anti-GFP antibody choices

A variety of anti-GFP antibodies are now provided to fluorescent protein users. In addition to the monoclonal anti-GFP antibody that has been introduced together with the GFP-Trap product group, Allele Biotech also has a strong anti-GFP polyclonal product, and a new rat anti-GFP monoclonal antibody. The availability of these different species specificity should allow users to have options in staining, detection, co-IP, double-labeling, etc.

New Product of the Week 04-05-10 to 04-11-10: Rat anti-AvGFP antibody, ACT-CM-MRGFP10

Promotion of the Week 04-05-10 to 04-11-10: Buy any Allele Biotech/Orbigen’s polyclonal antibody, get another of equal or less value at 60% off (if you are not a fan or friend through Allele’s online social networks, the discount is 30%).

Tags: 3H9, anti-GFP, antibodies, camelid antibodies, Chromotek, GFP booster, GFP detection, monoclonal, polyclonal, rat monoclonal

Using 2A “self-cleaving” peptide in bicistronic mammalian expression

Multiple promoters or internal ribosomal entry sites (IRES) have been used for the production of multiple proteins from the same vector. Potential drawbacks with multiple promoters on viral vectors include unstable genome and interference between promoters. IRES is a relatively large sequence that can cause problems in virus packaging, especially for viruses with very limited genome size such as AAV. In addition, it is required that the start of the second ORF is fairly close to the IRES, adding difficulties to cloning.

2A or 2A-like peptide (collectively called 2A peptide here) is used by several families of viruses, the best known foot-and-mouth disease virus of the Picornaviridae family, for producing multiple polypeptides. Although called a “self-cleaving” peptide or protease site, the mechanism by the 2A sequence for generating two proteins from one transcript is by ribosome skipping–a normal peptide bond is impaired at 2A, resulting in two discontinuous protein fragments from one translation even.

The 2A-based bicistronic expression has been used for several years, but recently gained much more popularity due to its successful use in iPSC generation that required 2 to 4 factors working in concert. Even expression of all factors can be achieved when 2A peptides are used for multiple protein production, due to near 100% efficiency of the 2A “cleavage” at each site, and no interference between multiple 2A sites. Early work used a 36 amino acid sequence as 2A peptide, which was later reduced to about half that size from mutation and screening. Commercial vectors utilizing 2A for co-expression of cDNA and fluorescent protein and/or drug resistance genes have not been available until now. Allele Biotech has introduced a number of such plasmids, establishing another First-to-the-market as it has done many times previously in its 10 year history.

New product of the week 03-29-10 to 04-04-10:

Alleleustrious pmTFP1-2A Bicistronic mammalian expression vector, ABP-FP-T2A10, $399, http://www.allelebiotech.com/shopcart/index.php?c=215&sc=34

Promotion of the week 03-29-10 to 04-04-10:

Buy any GFP-Trap beads or kits, get polyclonal anti-GFP (ABP-PAB-PAGFP10) at half size for FREE!  ***GFP-Trap has been replaced with GFP-nAb, an improved version for the same purposes in the form of agarose beads, magnetic beads, polyacrylamide beads, spin column kits, etc.  Come and take a look at the most comprehensive list of nearly all FPs before purchasing.

Tags: 2A peptide, anti-GFP, bicistronic, drug resistance, GFP-Trap, iPS, iPSCs, IRES, polycistronic, ribosome skip, self-cleaving, transcription, translation