Blogs for March 2017

Why you should use a Rabbit Monoclonal Antibody

Thursday, March 30, 2017 - 11:50

Primary antibodies are becoming increasingly popular investigate tool to use in research, given the breadth and selection of product on the market. Not only are antibodies raised in a variety of hosts, they also have proven reactivity with a wide range of species applying to many research fields and models. From here one can select a desired clonality, choosing polyclonal or monoclonal depending on the target of interest.  With monoclonal antibodies, it can be taken one step further to hone in on a specific clone of interest. With all of the different options available, it can be difficult to choose your ideal antibody.  Choosing the correct make up of an antibody will no doubt have an effect on your experimental outcomes,...

Potential therapies for human ovarian cancer surrounding the PI3K/AKT/mTOR pathway

Tuesday, March 28, 2017 - 09:47

Mammalian target of Rapamycin (mTOR) is a serine/threonine kinase that regulates cell growth, proliferation, motility and survival. Acting downstream of AKT, also a serine kinase, mTOR is composed of the mTORC1-Raptor complex and the mTORC2-Rictor complex.  mTOR has been implicated in many cancers, including ovarian cancer, due to the effect of mTOR inhibitors on tumor progression.  In addition, mTORC2 has a positive feedback effect on AKT behavior, which may explain its rapamycin resistance.  Rapamycin is a common drug used as a targeted therapy for mTOR inhibition, however it is not successful in all cancer types. In Ovarian Cancer specifically, the PI3K/AKT/mTOR pathway is the most frequently altered pathway, with PI3K, AKT and mTOR expression levels correlating with survival and tumor growth.  In the following articles, an mTOR antibody is used to investigate the efficacy of two...

What are the major differences between Apoptosis, Necroptosis & Autophagy?

Friday, March 24, 2017 - 13:41

Apoptosis is a form of programmed cell death which is mediated by cysteine proteases called caspases. It is an essential phenomenon in the maintenance of homeostasis and growth of tissues, and it also plays a critical role in immune response. The cytomorphological alterations and the key features of apoptosis are listed below:

apoptosis

...

Process

Active, physiological or pathophysiological

Induction stimuli

Oxidative stress, death receptor ligands, chemotherapy

Ultrasensitive IHC Detection with HRP-Polymer Conjugates

Thursday, March 23, 2017 - 14:54

Signal amplification methods are widely used in immunohistochemistry (IHC) for detection of rare epitopes and low abundance antigens. While many of these techniques such as the avidin-biotin complex (ABC) method improve staining, they frequently require additional steps and result in higher background staining.  Blocking endogenous biotin, a requirement of using ABC reagents, may not sufficiently remove residual activity in frozen tissue sections and tissues high in biotin including the liver and kidney.

FABP1/L-FABP antibody

Figure 1. FABP1/L-FABP was detected in paraffin-embedded sections of human kidney using Mouse Anti-Human FABP1/L-FABP Monoclonal Antibody (Catalog # MAB2964) at 1 μg/mL for 1 hour at room temperature followed by incubation for 30 minutes at room temperature with Anti-Mouse/Rabbit IgG VisUCyte HRP Polymer Antibody (Catalog...

The use of apoptosis antibodies and controls in cell death research

Monday, March 20, 2017 - 15:06

Apoptosis is a method of programmed cell death that is notably characterized by a morphological change in cellular nuclei and membrane appearance.  Not to be confused with necrosis, apoptosis is a pathway that is induced by a variety of factors that activate cysteine proteases known as caspases to lead the cell to its ultimate death versus natural death of a cell. While excessive apoptosis can be detrimental, it can also be part of our immune response or in protection of cells that have been damaged by disease or toxic stimuli.  Using antibodies against apoptotic pathway proteins is an effective way to investigate the role of apoptosis in a number of experimental models.  However, it is important to use antibodies against well-defined key apoptotic players and to integrate controls when possible. 

...

Frontiers in immunohistochemical (IHC) analysis

Thursday, March 16, 2017 - 09:06

There are a variety of experimental methods to choose from when using antibodies as a probe to highlight a target of interest. Western blot will reveal protein abundance and behavior, immunocytochemistry allows a look at protein behavior on the cellular level, and flow cytometry has the ability to label and sort hundreds of thousands of cells in no time. However, immunohistochemistry (IHC) is the only method where researchers have the ability to view the spatial localization of a target within a specific tissue. A lot has changed with IHC since its introduction in the mid 1900s, including automated processing machines, multiplexing techniques, advanced image analysis, robustly

Epidermal growth factor receptor (EGFR) in head and neck squamous cell carcinoma (HNSCC)

Tuesday, March 14, 2017 - 09:24

Head and neck squamous cell carcinoma is quite common in the U.S., covering more than 4% of all cancers each year, and is most susceptible to individuals between 50 and 60 years of age.  Squamous cells are a type of epithelial cell that are located all over the body with concentrations in the mouth, throat, neck and cervix.  EGFR, or epidermal growth factor receptor, is a trans-membrane glycoprotein that oversees cellular proliferation through its intrinsic tyrosine kinase activity.  When EGFR is bound to its ligand, it is phosphorylated by inner tyrosine kinase activity, where downstream pathways are activated and tumors caused by over stimulated cellular proliferation may occur.  Because of EGFR’s role in regulating multiple signaling cascades, and for the potential treatment therapies targeted at kinase inhibitor activity, EGFR is of high interest in understanding head and neck squamous cell carcinoma.  The following articles used an...

Topics in CD11b: The innate immune response

Thursday, March 9, 2017 - 11:58

Integrins are transmembrane receptors composed of alpha and beta chains, where beta-integrins are mainly expressed in leukocytes. Leukocytes are white blood cells that act in the immune system to defend our body against foreign pathogens.  Integrins are known to interact with extracellular matrix molecules to initiate our inflammatory immune response, in addition to regulating cell adhesion, migration and proliferation.  Our innate immune response is composed of a number of cell types that work in a coordinated effort to identify and attack foreign particles through antigen production.  CD11b is expressed in macrophages, monocytes, dendritic cells and granulocytes – the main players of our immune response cell network.  While CD11b’s exact role in immunity is still debated, it has been established that CD11b both negative and positively regulates our immune response, often times via a TLR ligand.  The following research articles use a...

The role of Parkin and autophagy in retinal pigment epithelial cell (RPE) degradation

Tuesday, March 7, 2017 - 13:49

The root of Parkinson’s disease (PD) points to a poorly regulated electron transport chain leading to mitochondrial damage, where many proteins need to work cohesively to ensure proper function.  The two key players of this pathway are PINK1, also known as PTEN or PARK6, and Parkin, also known as PARK2 - where PINK1 acts as an upstream effector of Parkin to regulate mitochondrial dynamics.  Mitochondria must maintain a healthy equilibrium and do so by undergoing a series of fission and fusion events.  The proteins Drp-1 and OPA-1, respectively, govern these events. While PINK1 and Parkin are directly involved in the progression of PD, their role in maintaining mitochondrial health has associated them with autophagy and mitophagy events in other models such as the loss of retinal pigment epithelial cell (RPE)...

Article Review: Ly6E/K Signaling to TGF-beta Promotes Breast Cancer Progression, Immune Escape, and Drug Resistance

Friday, March 3, 2017 - 14:52

Today in breast cancer research, scientists are focused on determining the cause, risk, diagnostic testing options and treatment of this devastating disease.  Of particular interest is identification of potential therapeutic targets that are known to contribute to the progression of breast cancer in order to develop treatments against these specific genes or proteins.  This article review summarizes research completed by AlHossiny et al regarding the role of Ly6E/K signaling and TGF-beta is the progression of cancer, immune escape and subsequent drug resistance.  Initial work by this group shed light on the role of mouse Ly6A in the regulation of TGF-beta, PTEN and the ERK/AKT signaling pathways to develop resistance to radiation and promote metastatic behavior of mammary tumors.  Furthermore, heightened levels...

The use of the autophagy marker LC3 in western blot, immunocytochemistry and flow cytometry research applications as an indicator of autophagy

Thursday, March 2, 2017 - 08:44

The process of autophagy, or lysosome-mediated degradation of damaged proteins and organelles in the cytosol, is a vital cellular process that acts as a quality control mechanism for proteins and organelles. The misregulation of autophagy can lead to an imbalance of cellular homeostasis and the subsequent development of disease.  Therefore, the study of autophagy is at the forefront of neuroscience and cancer research, among others.  In order to measure autophagic flux, many assays use the autophagy marker protein LC3.  LC3, the mammalian homolog of yeast ATG8, is a ubiquitin like protein that is associated with the autophagosome during the autophagic process. More directly related to LC3 is the process of selective autophagy, where receptors such as p62, NBR1 and NDP52 possess an LC3-interacting region (LIR) to directly bind LC3. Using a


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