Antibody News

Success of combined IL-10 and IL-12 therapy in colon cancer depends on IFN-gamma and gut barrier integrity

Tuesday, April 23, 2019 - 08:28
IL10 receptor alpha expression in human colon carcinoma, IHC

By Jamshed Arslan, Pharm. D., PhD.

Colon cancer is responsible for over 600,000 deaths per year worldwide. Colon cancer can be classified into two categories: mismatch repair (MMR)-deficient and MMR-proficient cancers.1 MMR-deficient colon cancer shows high CD8+ cytotoxic T-lymphocyte (CTL) and low Th17 cells, while MMR-proficient colon cancer shows an opposite trend. MMR-proficient colon cancer is associated with poor prognosis and is resistant to anti-cancerous immune checkpoint inhibitors. In other words, outcome of immune therapy is...

Visualizing Autophagy by Microscopy

Tuesday, April 16, 2019 - 09:36
LC3B  and alpha tubulin expression in HeLa cells treated with Chloroquine, ICC

By Christina Towers, PhD

Autophagy is a recycling process that relies on the formation of a unique organelle termed an autophagosome. An elegant way to monitor autophagy is through various microscopy techniques to visualize and quantify the formation and turnover of these double membrane structures. Some techniques are more quantitative than others, each accompanied by advantages as well as caveats.

Identifying the autophagosome with Transmission Electron Microscopy (TEM)

This technique is the only tool that can reveal all of the dynamic stages of a forming autophagosome in the nanometer range and in its...

How to identify B cell subsets using flow cytometry

Tuesday, April 9, 2019 - 08:49
Explore new Flow Cytometry Handbook, Bio-Techne

By Victoria Osinski

Using flow cytometry to identify B cell subsets

Identifying cellular subsets by flow cytometry requires careful and thorough planning in order to ensure the correct subset of cells are identified and isolated. While generally thought of as antibody-secreting cells, an expanding body of research proves that B cells are a heterogeneous population. Flow cytometry can be used to identify many of these B cell subsets, so long as important nuances and technical approaches are considered.

Setting up to identify B cell subsets

Before subsetting B cells, steps must be taken to ensure dead cells, cell aggregates, and debris are not...

Lipopolysaccharide from gut microbiome localizes in human atherosclerotic plaques and promotes TLR4-mediated oxidative stress

Tuesday, March 26, 2019 - 09:05
TLR signaling pathway, Novus Biologicals

By Jamshed Arslan, Pharm. D., PhD.

Atherosclerosis is a chronic inflammatory condition in which plaques of fats and other substances slowly buildup on the inner walls of arteries to restrict blood flow. In atherosclerosis-associated ailments like obesity, hypertension and type II diabetes, intercellular tight junctions are impaired, leading to increased gut permeability. This means that products of intestinal microbiota like lipopolysaccharide from Gram-negative bacteria can seep into the blood stream.

A pro-atherogenic role of lipopolysaccharide has been suggested. For example,...

Toll-like receptors in the intestinal epithelial cells

Tuesday, March 19, 2019 - 10:30
Defensin alpha 5 expression in human small intestine, IHC

By Jamshed Arslan, Pharm. D., PhD.

Toll-like receptors (TLRs) are microbe-sensing proteins that act as first responders to danger signals. TLRs help the intestinal epithelial cells (IECs) recognize commensal bacteria from the harmful/foreign ones. Polymorphisms and variants of TLRs have been linked with immune system’s attack on commensal bacteria, leading to inflammatory bowel disease and colorectal cancers. The endogenous pattern of TLR expression in IECs and the functional outcome of TLR signaling in vivo are not well defined due to various technical difficulties. Thanks to the researchers in institutes in California and...

Probiotics and exercise can slow Alzheimer’s progression by altering composition of gut bacteria

Tuesday, March 12, 2019 - 09:11
beta amyloid expression in human Alzheimer’s disease hippocampus, IHC

By Jamshed Arslan, Pharm. D., PhD.

Gut microbiome modulates various disorders even outside the gut. For example, the microbiome is linked to amyloid-β deposition and the consequent inflammation in Alzheimer’s dementia. Altering gut microbiome through probiotics and/or exercise has been hypothesized to improve memory in Alzheimer’s disease (AD). A multi-continental collaboration of scientists tested this hypothesis on AD mice and found that probiotics and exercise can retard AD progression, at least partly by changing the microbiome.

Probiotics with exercise improve memory in AD mice

The researchers used Framelim, a product...

Microglia: pruning shears for homeostatic maintenance in the brain

Tuesday, March 5, 2019 - 08:58
Iba1 expression in mouse fore brain, IHC

By Jennifer Sokolowski, MD, PhD.

Microglia play a critical role in pruning neurons and synapses during homeostatic maintenance in the adult brain.1 A recent study by Ayata et al. (2018) identified regional differences in clearance and found a role for epigenetic control via polycomb repressive complex 2 (PRC2) in adult rodents.2

How to evaluate microglial clearance?

Ayata et al. defined a cell clearance phenotype by labeling microglia with iba1 and CD68 and evaluating morphology, cell volume, and CD68+ lysosome content. They also...

The LC3 A, B, C’s and 1, 2, 3’s

Tuesday, February 26, 2019 - 13:38
Atg5 expression in HeLa cells, ICC

By Christina Towers, PhD

Autophagy is a catabolic process used to breakdown and recycle damaged proteins and organelles. It is a multistep process that, in its simplest form, consists of 4 steps: initiation, phagophore nucleation, elongation, and fusion – resulting in a double membrane structure that upon fusion with the acidic lysosome, facilitates the degradation of its contents. LC3B (and its family members) are incorporated into the elongating autophagophore and are necessary for efficient autophagosome formation and autophagic degradation. During lysosomal fusion, LC3B is itself degraded. For these reasons, LC3B protein levels are often...

Best way to quantitatively measure Autophagic Flux

Tuesday, February 19, 2019 - 10:36
LC3B expression in HeLa cells, ICC

By Christina Towers, PhD

Autophagy is a stress induced cellular recycling process that plays an important physiological role in many diseases. It is induced by a variety of stimuli both intracellular and extracellular and is a tightly regulated process with over 30 core autophagy proteins necessary for efficient autophagosome formation and fusion with lysosomes to facilitate degradation1. One of the most common ways to monitor autophagy is by measuring the protein levels of LC3-II, which is incorporated into autophagosomes and then degraded in the...

Wet or dry protein transfer: Which one to choose for your western blotting?

Tuesday, February 12, 2019 - 10:50
Novus Biologicals western blot resources page

By Jamshed Arslan, Pharm. D., PhD.

Western blotting involves three steps: 1) separating proteins by size through gel electrophoresis 2) transferring separated proteins to a solid support (typically, a PVDF membrane) and 3) using antibody to mark your protein(s) of interest in the mixture of proteins. The second step of moving negatively charged proteins from the gel to a blotting membrane (PVDF) can be performed in dry or wet conditions.

Dry/semi-dry transfers

Dry conditions are famous for easiness and speed. Reagents, power packs and wiring involved in dry transfers are less messy and therefore, potentially less hazardous. They allow blotting of several...

Animal Models to Study Autophagy

Tuesday, February 5, 2019 - 09:03
Autophagy Signaling Pathway, Novus Biologicals

By Christina Towers, PhD

What is autophagy?

Autophagy is the catabolic process that degrades cytoplasmic material via the lysosome. In the beginning... the process of macroautophagy was originally characterized in yeast where the core autophagy proteins or Apgs were identified. In this first system, the multistep process of phagophore initiation and elongation as well as the conjugation machinery that mediate efficient autophagosome completion were determined and cloned to assess function. The mechanisms of starvation induced autophagy as well as many of the selective autophagy processes were also hashed out in...

Sample collection from mammalian culture cells for kinomic analysis

Tuesday, January 29, 2019 - 10:10
ERK1/2/5 Signaling Interactive Pathway

By Jamshed Arslan Pharm.D., PhD.


Kinome describes kinases, and kinomics refers to the kinase signaling. Studying the effects of reagent (exogenously applied growth factor or inhibitor) on kinase activity is a common experiment in cell, developmental and integrative biology laboratories. This blog offers a cost-effective and robust method of obtaining adequate amount of sample for kinomic analysis from mammalian culture cells like HEK cells after treatment with reagent. Using this method, you can easily get over 50 μg protein sample in 50 μl volume from a confluent well of a standard 6-well plate with 0.5 x 106...

Best Methods to Induce and Inhibit Autophagy Pharmacologically

Tuesday, January 22, 2019 - 09:33
VPS35 expression in endosomes and lysosomes of human U-2 OS cells, ICC

By Christina Towers, PhD

Autophagy facilitates the degradation and recycling of damaged cytoplasmic material. The multistep process consists of a double membrane structure called an autophagosome that engulfs proteins and organelles and then fuses with lysosomes to facilitate their breakdown. Autophagy is tightly regulated by nutrient availability and is negatively regulated by the robust nutrient sensor mammalian target of rapamycin (mTOR). Starvation, either by glucose deprivation or amino acid depletion, will cause a marked increase in autophagy, best quantified by an increase in autophagic flux. Likewise,...

Phagocytes in Multiple Sclerosis: Myelin uptake leads to oxysterol-induced activation of liver X receptors, LXRs

Tuesday, January 15, 2019 - 08:41
Myelin basic protein (MBP) expression in cerebellum tissue IHC

By Jamshed Arslan Pharm.D., PhD.

Myelin is a cholesterol-rich layer around nerves. Damage to this protective layer and the consequent slowing down of nerve impulses are hallmarks of multiple sclerosis (MS) and other demyelinating diseases. Myelin breakdown alters plasma levels of 27-hydroxycholesterol (27OHC) and other CNS-specific oxysterols, which are natural ligands of transcription factors called liver X receptors (LXRs). Since myelin phagocytosis is a source of intracellular oxysterols in the resident microglia and infiltrating macrophages, a team of scientists in Europe decided to test the hypothesis that myelin uptake and subsequent...

mTOR Signaling and the Tumor Microenvironment

Tuesday, January 8, 2019 - 09:10
mTOR signaling pathway poster

By Yoskaly Lazo-Fernandez, PhD

The mammalian target of rapamycin (mTOR) is a conserved serine/threonine kinase that, as a member of two distinct intracellular protein complexes, mTORC1 and mTORC2, regulates protein synthesis, autophagy metabolism, proliferation and survival. More and more research is showing how the catalytic activity of both mTOR complexes plays an important role in cancer biology. Not only mTOR is upregulated and contributes to the development of numerous cancer types, but it also contributes to the angiogenic and immune responses within the tumor microenvironment (TME)1.


NOX2 oxidase is a potential target for broad-spectrum antiviral therapies

Wednesday, January 2, 2019 - 09:24
TLRs signaling pathway – interactive

By Beth Melson, MS

Viral infection induces the production of reactive oxygen species (ROS) by inflammatory cells, the majority of which is produced by the NOX2 oxidase. While NOX2 oxidase contributes to the clearance of bacteria and fungi, its role in viral infection is less clear, and in fact seems to contribute to increased viral pathogenicity. Recently, it was demonstrated that viruses, including influenza A, induce NOX2 oxidase production through suppression of TLR7 sensing single-stranded RNA (ssRNA)1. Therefore, inhibition of the NOX2 oxidase pathway represents an opportunity for viral...

TMEM 119 is a specific marker of microglia cells

Thursday, December 20, 2018 - 08:58
P2Y12/P2RY12 expression, human cerebral cortex shows cytoplasmic positivity in microglia, IHC

By Jennifer Sokolowski, MD, PhD.

Microglia are a major immune-cell component in the brain. They ingest and degrade dead cells, debris, and foreign material and interact with other immune cells to orchestrate central nervous system immune responses.1,2 Microglia appear to play a critical role in modulating normal physiologic immune functions as well as the immune response in disease states. In order to define the role of microglia, we need specific markers that allow distinction of microglia from other cell types; fortunately, researchers have validated a new microglia-specific marker,...

The role of Wnts in neuroinflammation

Tuesday, December 18, 2018 - 08:35
TRPM2 expression in rat hippocampus neurons, IHC

By Michalina Hanzel, PhD

The multifaceted roles of the Wnt family of glycoproteins have been extensively characterized throughout embryonic development and adult homeostasis. The highly conserved, cell- and tissue- specific proteins orchestrate processes ranging from neural induction, cell proliferation and migration to adult neurogenesis and neuronal maintenance and regeneration. Recently, Wnt proteins have been proposed to have key roles in regulating pathological processes in the brain, especially those observed in neurodegenerative disorders, often resulting from pathological states of neuroinflammation.


RNA-binding protein Staufen1 conspires with Atxn2 in stress granules to cause neurodegeneration by dysregulating RNA metabolism

Tuesday, December 11, 2018 - 11:19
PCP4 expression in mouse cerebellum Purkinje neurons, IHC

By Jamshed Arslan Pharm.D.

Spinocerebellar ataxia type 2 (SCA2) is a movement disorder characterized by neurodegeneration. The cause of this autosomal dominant disease is a mutation in the RNA processing gene Atxn2, which leads to polyglutamine (polyQ) expansion in ATXN2 protein. The association between mutant polyQ proteins, neurodegeneration, and dysfunctional autophagy has been extensively studied in amyotrophic lateral sclerosis (ALS), autism, and Huntington’s disease, but not in SCA2. It is noteworthy that ATXN2 is widely expressed in our nervous system, partly as a constituent of stress granules (SGs), which...

Flow Cytometry Basics for the Non-Expert

Tuesday, December 4, 2018 - 10:35
Flow cytometry dot plots, histograms, density plots

By Jody Bonnevier, PhD.

Flow cytometry: A Challenging Interdisciplinary Application

Flow cytometry has long been the cell analysis method of choice for immunologists, however more and more disciplines have recognized its powerful and flexible capabilities, and want to harness the advantages of this multiparameter single-cell analytical tool. Nowadays, the benefits of flow cytometry for cell phenotyping extend from basic to clinical research and diagnostic applications as well as to drug discovery and development.

In basic research, the same flow cytometry techniques used to phenotype T cell subsets can be applied to defining protein...

Losing memory: Toxicity from mutant APP and amyloid beta explain the hippocampal neuronal damage in Alzheimer's disease

Tuesday, November 27, 2018 - 13:42
Map2 and Fox3/NeuN expression in rat hippocampus, IHC

By Jamshed Arslan Pharm.D.

Alzheimer's disease (AD) is an irreversible brain disorder that destroys memory and thinking skills. The telltale signs of AD brains are extracellular deposits of amyloid beta (a polypeptide that comes from proteolysis of precursor protein APP) and intracellular neurofibrillary tangles. The consequent neurodegeneration affects regions associated with memory and cognition, the most noteworthy of which is the hippocampus. It goes without saying that generating immortalized primary hippocampal neurons that could express AD-related protein(s) would immensely...

Application Focus: I see an increase in LC3, now what?

Monday, November 19, 2018 - 13:05
LC3A expression in HeLa cells, chloroquine treated, ICC

By Christina Towers, PhD.

Autophagy is highly conserved and tightly regulated process that all cell types use to recycle nutrients, particularly in the instance of stress1. As a result, even small changes in signaling pathways, gene expression, or drug treatments can alter autophagy. If you stumble into the field of autophagy by happenstance and hypothesize a change in autophagy with your favorite cell manipulation of choice, you probably read a handful of publications and came to the conclusion that you should assay LC3 levels to test this hypothesis. In fact, LC3 expression can be a good readout for autophagy, but researchers should be...

E-syt in Autophagosome biogenesis: What is the source of it all?

Tuesday, November 13, 2018 - 13:44
Calreticulin expression in vesicles and endoplasmic reticulum of HeLa cells, ICC

By Christina Towers, PhD.

Macroautophagy is a cellular recycling process that requires the formation of double membrane structures to engulf and degrade damaged cytoplasmic material. The pathway involves over 20 core autophagy proteins (

Negative feedback regulation of EPAS1 gene in non-small cell lung cancer through DNA methyltransferases

Tuesday, November 6, 2018 - 09:29
HIF-1 alpha expression in human lung tissue, IHC

By Jamshed Arslan Pharm.D.

Non-small cell lung cancer (NSCLC) is the most common type of lung cancer. NSCLC cells show increased expression of HIF-1alpha and HIF-2 alpha, which help cells overcome hypoxia. However, NSCLC samples have shown decreased mRNA levels of HIF-2 alpha-encoding gene called EPAS1. The mechanism of downregulation of EPAS1 transcription in NSCLC is unknown. A team of researchers from various institutes in Shanghai and Yangzhou, China, suspected transcriptional inactivation through DNA...

Nickel induces migratory and invasive phenotype in human epithelial cells by epigenetically activating ZEB1

Tuesday, October 30, 2018 - 09:05
Epigenetic mechanisms Novus Biologicals

By Jamshed Arslan Pharm.D.

Nickel (Ni) is a naturally abundant metallic element. It is a major component of stainless steel, coins, and many other items of daily use. Disturbingly, Ni exposure is associated with cancer and several diseases of the lung, kidney and cardiovascular system. Despite being carcinogenic, Ni’s ability to cause DNA mutations and induce oxidative stress is low. To explain this paradox and investigate how the impact of carcinogen exposure may last for months or even years, researchers at the New York University and University of Virginia examined Ni-induced changes in transcription and cellular regulation in human epithelial cells from lung and urinary...


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