Sensitive, Specific and Cost Effective Protein Expression Analysis

Reaching for Cell Based Assay Solutions Excellence

We have been receiving positive feedback from clients using our Quantibody® and RayBio® C-Series Membrane-Based Antibody Arrays. Our manufacturing partner, Raybiotech, is ISO 9000 certified insuring all arrays are made to exacting specification. The cost for measuring a single cytokine runs as low as 16 USD/Cytokine!

We use these arrays to test the blood serum of clients with autoimmune diseases to determine levels of cytokine and growth factor expression vs healthy controls. We also use them to determine proteins secreted by our  USB Derived hMSCs cultured in our hMSC Media. This gives us insight as to the protein expression patterns in our hMSC, Chondrogenesis and Osteogenesis Assays.

Figure: The graph shows cytokine secretion of hMSCs after a continuous 24 day culture. These results were obtained using our select markers from our cytokine and bone metabolism arrays. Data courtesy of Dr. Jim Musick and Tiana Tonrey, Vitro Biopharma.

We are pleased to announce the addition of these new human and mouse arrays:

Quantibody® Human Growth Factor+Neurotrophin+Cytokine Array Q1-10. Precisely measures: Amphiregulin, BDNF, bFGF, BMP-4, BMP-5, BMP-7, beta-NGF, EGF, EGFR, EG-VEGF (PK1), FGF-4,FGF-7 (KGF), GDF-15, GDNF, Growth Hormone, HB-EGF, HGF, IGFBP-1, IGFBP-2, IGFBP-3, IGFBP-4, IGFBP-6, IGF-1, Insulin, M-CSF R, NGFR (TNFRSF16), NT-3, NT-4, Osteoprotegerin (TNFRSF11B), PDGF-AA, PLGF, SCF, SCF R (CD117/c-kit), TGF alpha, TGF beta 1, TGF beta 3, VEGF-A, VEGFR2, VEGFR3 and VEGF-D.
Mouse C-Series Cytokine Array. C1-2-Semi-quantitavely measures: GCSF,GM-CSF, IL-2, IL-3, IL-4, IL-5, IL-6, IL-6, IL-9, IL-10, IL-12 p40/p70, IL-12 p70, IL-13, IL-17A, IFN-gamma, MCP-1 (CCL2), MCP-5, RANTES (CCL5), SCF, TNF RI (TNFRSF1A), TNF alpha, Thrombopoietin, (TPO) and VEGF-A.

Images: Representative images obtained with RayBio® C-Series Antibody Arrays. These membranes were probed with conditioned media from two different cell lines. Membranes were exposed with Kodak X-Omat® film at room temperature for 1 minute. Note the strong signals of the Positive Control spots in the upper left and lower right corners.

Questions? Do not hesitate to contact me: pshuster@neuromics.com or direct phone-612-801-1007. Thank you. Pete Shuster, CEO and Owner, Neuromics

Cytokines and Neuropathic Pain

I have been frequently posting results of testing blood serum of our clients suffering from Autoimmune (including Neuroimmuno) Diseases and Autistic Children. Our testing includes both standard medical testing (serum levels of toxins, metals, pathogen loads and markers like Cortisol, CRP, CDs, ALPS, NSE, S100b). In addition, we also doing testing using our Quantibody® Antibody.

Many of the clients report chronic mild to severe neuropathic pain.

By definition, this pain encompasses a series of heterogeneous conditions with some similar clinical manifestations. Peripheral examples include traumatic nerve injury, diabetic peripheral neuropathy and chemotherapy-induced peripheral neuropathy or Multiple Sclerosis. (see: Front Pharmacol. 2013; 4: 142.Published online Nov 22, 2013. doi: 10.3389/fphar.2013.00142). It is hard to unravel the cause and effects of this pain,

However, in analyzing the results from our clients' test we notice a relationship between dysregulation of some key cytokines and levels of pain (more data is required to confirm this). Here're our results (n=5)

Figure: Clients reporting pain vs healthy controls.

We will continue posting results here. We are releasing a Custom Quantibody Neuroimmuno-Pain Array which will include more bio-markers suspected to play a role in pain signaling.

Autism Spectrum Disorder (ASD) Custom Array Results

Blood Serum Levels and Key Markers

We have been testing immune/inflammatory response, oxidative stress and growth factor markers in ASD children from central Europe. Most showed high levels of  related cytokines and chemokines.

From our initial testing and published results, we have developed a custom Quantibody Array to test ASD children. The markers in this array are: BDNF, HSP-70, Leptin, RAGE and TGF-beta1.

Figure: Serum Levels of Key Markers in Tested ASD Children

Here's more on each marker:

• BDNF is a protein involved in making healthy new neurons. This protein is dis-regulated in autism. It is shown to be decreased in some studies and increased in others. These variations could be a function of age. During brain development, BDNF regulates the birth and differentiation of brain cells, or neurons. Some of BDNF’s target cells, such as cortical interneurons, which transmit information between different layers of the brain cortex, have been implicated in autism. BDNF is also a regulator of brain growth, and children with the disorder tend to have abnormally large brains during early development. Vigorous exercise, for example, increases BDNF levels in blood and studies have linked this increase to growth of new healthy neurons in the hippocampus region of the brain. We found all ASD children tested showed low levels of BDNF. All had moderate to high levels of heavy metals and viral/bacterial pathogens. Most also had evidence of leaky blood brain barriers. Could this low level be due to consumption of BDNF demanded by the chronic need for neuro-repair or is genetic in origin or perhaps both?
• HSP70 is strongly upregulated by heat stress and toxic chemicals, particularly heavy metals such as: aluminum, arsenic, cadmium, copper, mercury, etc. This upregulation in ASD could be linked to difficulty is in clearing toxins. Children with highest tested HSP70 levels also had the highest concentration of heavy metals. The 3 children with healthy levels have been undergoing ongoing treatment to clear metals.
• Leptin modulates appetite and energy tough elevated levels of leptin present in cases of autism might be an important sign of immune processes, particularly those related to inflammation. It is also suggested leptin may be a link between autism and epilepsy that provides an avenue for novel or better management of autistic children with epilepsy. All were in the range of published healthy controls (mean=2065 pg/ml).
• RAGE is hypothesised to have a causative effect in a range of inflammatory diseases such as diabetic complications, Alzheimer's Disease and even some tumors. In ASD, it is thought to be a master switch for chronic inflammation. Including in the brain. RAGE is a receptor for S100B so it is also elevated in ASD and is reflective of neurological damage. All showed elevated serum levels of rage.
• TGF-beta1 is a protein that controls proliferation, cellular differentiation, and other functions in most cells. This control includes tissue repair in the nervous system. Decreased transforming growth factor beta1 in autism is a potential link between immune dysregulation and impairment in clinical behavioral outcomes. 6 of 8 of the children had low TGF-beta1 levels. Though the correlation between symptoms and behaviors is not conclusive.

We plan to significantly increase the samples of ASD children tested using this custom Quantibody Array. . 

Our ultimate goal is to use this array as a tool to determine the efficacy of therapies we are developing. Our goals is to develop natural products based therapies that are proven to activate stem cells. These cells could catalyze immune response modulation and tissue repair. The process is to first clear metals and pathogens, then treat with stem cell activators and test and fine tune treatments and test again. 

I will be posting results on an ongoing basis. I also welcome any and all comments. I am available for direct contact at pshuster@neuromics.com or 612-801-1007.

Quantibody® Cytokine Arrays

Cytokines, Adipokines, Growth Factors and Neurotrophins

We are pleased to announce addition of Antibody Arrays to our catalog. These are excellent tools for study apoptosis, inflammation, angiogenesis and  immune response and related autoimmune, inflammatory and degenerative diseases plus many forms of cancers. Check out this video.

Specific Research Applications Include:
•High-throughput profiling of cytokine expression
•Validation of semi-quantitative antibody array results
•Identifying potential molecular targets for drug development
•Identifying the molecular mechanisms of drug action
•Identifying crucial factors involved in disease processes
•Discovering biomarkers for disease management
•Discovering expression patterns for molecular classification of diseases

How Quantibody Arrays Work

We will be aggresively adding new arrays to our product offerings in June. Stay tuned for more.





STEMEZ(TM) hNP1 Cells and Neuroprotection Studies

Neuroinflammation has been shown to be a potent negative regulator of stem cell and progenitor cell proliferation in the neurogenic regions of the brain.

Here researchers used our STEMEZ(TM) hNP1 Human Neural Progenitors Expansion Kit to study the neuroprotection capabilities of a propriety nutraceutical formulation.

Adam D. Bachstetter, Jennifer Jernberg, Andrea Schlunk, Jennifer L. Vila, Charles Hudson, Michael J. Cole, R. Douglas Shytle, Jun Tan, Paul R. Sanberg, Cyndy D. Sanberg, Cesario Borlongan, Yuji Kaneko, Naoki Tajiri, Carmelina Gemma, Paula C. Bickford. Spirulina Promotes Stem Cell Genesis and Protects against LPS Induced Declines in Neural Stem Cell Proliferation. PLoS ONE 5(5): e10496. doi:10.1371/journal.pone.0010496.

Abstract:Adult stem cells are present in many tissues including, skin, muscle, adipose, bone marrow, and in the brain. Neuroinflammation has been shown to be a potent negative regulator of stem cell and progenitor cell proliferation in the neurogenic regions of the brain. Recently we demonstrated that decreasing a key neuroinflammatory cytokine IL-1β in the hippocampus of aged rats reversed the age-related cognitive decline and increased neurogenesis in the age rats. We also have found that nutraceuticals have the potential to reduce neuroinflammation, and decrease oxidative stress. The objectives of this study were to determine if spirulina could protect the proliferative potential of hippocampal neural progenitor cells from an acute systemic inflammatory insult of lipopolysaccharide (LPS). To this end, young rats were fed for 30 days a control diet or a diet supplemented with 0.1% spirulina. On day 28 the rats were given a single i.p. injection of LPS (1 mg/kg). The following day the rats were injected with BrdU (50 mg/kg b.i.d. i.p.) and were sacrificed 24 hours after the first injection of BrdU. Quantification of the BrdU positive cells in the subgranular zone of the dentate gyrus demonstrated a decrease in proliferation of the stem/progenitor cells in the hippocampus as a result of the LPS insult. Furthermore, the diet supplemented with spirulina was able to negate the LPS induced decrease in stem/progenitor cell proliferation. In a second set of studies we examined the effects of spirulina either alone or in combination with a proprietary formulation (NT-020) of blueberry, green tea, vitamin D3 and carnosine on the function of bone marrow and CD34+ cells in vitro. Spirulina had small effects on its own and more than additive effects in combination with NT-020 to promote mitochondrial respiration and/or proliferation of these cells in culture. When examined on neural stem cells in culture spirulina increased proliferation at baseline and protected against the negative influence of TNFα to reduce neural stem cell proliferation. These results support the hypothesis that a diet enriched with spirulina and other nutraceuticals may help protect the stem/progenitor cells from insults. Figure 7. Spirulina increases proliferation of human neural stem cells in vitro and protects against a TNFα insult.

Figure: Human neural progenitors grown under proliferation conditions were assessed by MTT assay (A) or BrdU (B) for the effects of spirulina (125 ng/ml) or NT-020 (500 ng/ml) or the two treatments combined in the presence or absence of TNFα (20 ng/ml) for 72 hours. (A) The MTT assay shows that spirulina alone or NT-020 alone increase proliferation; surprising, the in combination proliferation is decrease compare to control ** p less tan 0.005.

Related Reagents:

STEMEZ(TM) hNP1 Human Neural
Progenitors Expansion Kit

Primary Neurons and Astrocytes-Primary
human, rat and mouse neurons and astrocytes

Neuron/Glial Marker Antibodies

Neurotrophins and Growth Factor
Antibodies

Stem Cell Research Reagents-includes
cells, antibodies, proteins, media and FACS kits.