References

Ancheta L, Bouajram R, Lappi DA (2018) Screening targeting agents and their cell surface biomarkers for high specificity and rapid internalization via cell death and fluorescence. Neuroscience 2018 Abstracts 128.20 / M17. Society for Neuroscience, San Diego, CA.

Summary: Some of the most recent successes in the treatment of cancers or research into passive immunotherapies for neurodegenerative diseases, employ the use of antibodies. These treatments utilize antibodies that either: 1) interfere with cell surface proteins responsible for tumor cell proliferation, 2) act as immune checkpoint inhibitors, or 3) are re-engineered to allow transport of other molecules across the blood-brain barrier (BBB). There are a growing number of antibody and small molecule therapeutic candidates and this demands a quick and efficient technique to screen for biomarkers that internalize effectively upon binding. The method described provides for the efficient determination of internalization of cell surface biomarkers upon binding of antibodies or peptides. This one-step, robust method uses a targeting agent combined with both a fluorescent reporter and a cytotoxic payload. The construct that makes this method effective was formed by cross-linking a fluorescent reporter, in this case fluorescein (FITC) and streptavidin to the ribosome-inactivating protein, Saporin. The conjugate used in screening potential therapeutics is a mixture of a biotinylated targeting agent mixed in a 1:1 molar ratio with FITC-labeled Streptavidinylated-Saporin. The method provides a definitive assay readout: fluorescence within 1 hour and cell death in 72 hours. This method is designed for rapid screening, in a quick and reproducible manner, for specificity and internalization in various cell types to explore suitability of candidates as therapeutics.

Related Products: Streptavidin-ZAP (Cat. #IT-27), FITC-Streptavidin-ZAP (Cat. #IT-85)

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See Also:

• Tan HL et al. Conservation of oncofetal antigens on human embryonic stem cells enables discovery of monoclonal antibodies against cancer. Sci Rep 8:11608, 2018.
• Yuan X et al. Characterization of the first fully macropinocytosing human antibodies human anti-TEM1 scFv in models of solid tumor imaging and immunotoxin-based therapy. Cancer Immunol Immunother 66:367-378, 2017.
• Forsyth PA et al. p75 neurotrophin receptor cleavage by α- and γ-secretases is required for neurotrophin-mediated proliferation of brain tumor-initiating cells. J Biol Chem 289(12):8067-8085, 2014.
• Thakkar JP et al. Epidemiologic and molecular prognostic review of glioblastoma. Cancer Epidemiol Biomarkers Prev 23(10):1985-1996, 2014.
• Kohls M Evaluate Potential Targeting Molecules. Nature Methods , 2006.

Zoccal DB, Silva JN, Barnett WH, Lemes EV, Falquetto B, Colombari E, Molkov YI, Moreira TS, Takakura AC (2018) Interaction between the retrotrapezoid nucleus and the parafacial respiratory group to regulate active expiration and sympathetic activity in rats. Am J Physiol Lung Cell Mol Physiol 315(5):L891-L909. doi: 10.1152/ajplung.00011.2018

Objective: To investigate the microcircuitry responsible for the distribution of the excitatory signals to the the parafacial respiratory group (pFRG) and the the respiratory central pattern generator (rCPG) in conditions of high CO2.

Summary: The activation of the pFRG late-E neurons during hypercapnia require glutamatergic inputs from the RTN neurons that intrinsically detect changes in CO2/pH.

Usage: Bilateral injections of SSP-SAP (0.6 ng in 100 nL of saline per side).

Related Products: SSP-SAP (Cat. #IT-11)

Qian L, Milne MR, Shepheard S, Rogers ML, Medeiros R, Coulson EJ (2019) Removal of p75 neurotrophin receptor expression from cholinergic basal forebrain neurons reduces amyloid-β plaque deposition and cognitive impairment in aged APP/PS1 mice. Mol Neurobiol 56(7):4639-4652. doi: 10.1007/s12035-018-1404-2

Objective: To investigate the contribution of CBF neuronal p75NTR to the progression of Alzheimer’s Disease

Summary: Data indicate that a direct interaction between CBF-expressed p75NTR and Aβ does not contribute significantly to the regulation of Aβ load.

Usage: To lesion CBF neurons, a single infusion of mu p75-SAP or control Rabbit IgG-SAP (0.4 mg/ml) was stereotaxically-injected into the basal forebrain.

Related Products: mu p75-SAP (Cat. #IT-16), Rabbit IgG-SAP (Cat. #IT-35)

Garrett AM, Khalil A, Walton DO, Burgess RW (2018) DSCAM promotes self-avoidance in the developing mouse retina by masking the functions of cadherin superfamily members. Proc Natl Acad Sci U S A 115:E10216-E10224. doi: 10.1073/pnas.1809430115

Summary: Focus on DSCAM (Down syndrome cell adhesion molecule 1) self-avoidance function in the mouse retina. DSCAM and members of the cadherin superfamily have also emerged as key contributors to a variety of neurodevelopmental disorders, including autism, schizophrenia, bipolar disease, Down syndrome and intellectual disability.

Related Products: Melanopsin-SAP (Cat. #IT-44)

Han W, Tellez LA, Perkins MH, Perez IO, Qu T, Ferreira J, Ferreira TL, Quinn D, Liu Z-W, Gao X-B, Kaelberer MM, Bohórquez DV, Shammah-Lagnado SJ, de Lartigue G, de Araujo IE (2018) A neural circuit for gut-induced reward. Cell 175:665-678. doi: 10.1016/j.cell.2018.08.049

Objective: To determine relevant gut-brain neuronal circuitry to motivational and emotional states.

Summary: There is a critical role for the vagal gut-to-brain axis in motivation and reward.

Usage: Injected 0.5 µl of CCK-SAP (250 ng/µl) into the R-NG of VGlut2-ires-Cre mice.

Related Products: CCK-SAP (Cat. #IT-31)

Acharjee S, Verbeek M, Gomez CD, Bisht K, Lee B, Benoit L, Sharkey KA, Benediktsson A, Tremblay M-E, Pittman QJ (2018) Reduced microglial activity and enhanced glutamate transmission in the basolateral amygdala in early CNS autoimmunity. J Neurosci 38:9019-9033. doi: 10.1523/JNEUROSCI.0398-18.2018

Objective: To identify CNS changes associated with behaviors in multiple sclerosis (MS) patients.

Summary: The data from this study reveal increased synaptic activity and spine density in early stages of experimental autoimmune encephalomyelitis (an animal model of MS) in the basolateral amygdala.

Usage: Mac-1-SAP mouse/human or Rat-IgG-SAP (control) was injected unilaterally in the BLA (1 ug/1 ul).

Related Products: Mac-1-SAP mouse/human (Cat. #IT-06), Rat IgG-SAP (Cat. #IT-17)

Lind LA, Murphy ER, Lever TE, Nichols NL (2018) Hypoglossal motor neuron death via intralingual CTB-saporin (CTB-SAP) injections mimic aspects of amyotrophic lateral sclerosis (ALS) related to dysphagia. Neuroscience 390:303-316. doi: 10.1016/j.neuroscience.2018.08.026

Objective: Despite its fundamental importance, dysphagia (difficulty swallowing) and strategies to preserve swallowing function have seldom been studied in ALS models.

Summary: The authors report a novel experimental model using intralingual injections of cholera toxin B conjugated to saporin (CTB-SAP) to study the impact of only hypoglossal motor neuron death without the many complications that are present in ALS models.

Usage: Hypoglossal motor neuron survival, swallowing function, and hypoglossal motor output were assessed in Sprague Dawley rats after intralingual injection of either CTB-SAP (25 ug) or unconjugated CTB and SAP (controls) into the genioglossus muscle.

Related Products: CTB-SAP (Cat. #IT-14)

Ebadi R, Kordi-Tamandani DM, Ghaedi K, Nasr-Esfahani MH (2018) Comparison of two different media for maturation rate of neural progenitor cells to neuronal and glial cells emphasizing on expression of neurotrophins and their respective receptors. 45(6):2377-2391. doi: 10.1007/s11033-018-4404-4 PMID: 30306506

Objective: To compare neural differentiation potential of two different media, NB + 5%ES-FBS + N2B27 and Ko-DMEM + 5%ES-FBS for conversion of mESC derived neural progenitors (NPs) into mature neural cells with emphasis on effect of these two media on neurotrophins and their respective receptors expression.

Summary: Results indicated that NB + 5%ES-FBS + N2B27 medium promoted neural differentiation process of mESCs and caused enhancement of neurotrophins protein expression in addition to their cognate receptors.

Usage: Western Blot (1:4000)

Related Products: NGFR (mu p75) Rabbit Polyclonal, affinity-purified (Cat. #AB-N01AP)

Mao S, Wang L, Chen P, Lan Y, Guo R, Zhang M (2018) Nanoparticle-mediated delivery of Tanshinone IIA reduces adverse cardiac remodeling following myocardial infarctions in a mice model: Role of NF-κB pathway. Artif Cells Nanomed Biotechnol 46(sup3):S707-S716. doi: 10.1080/21691401.2018.1508028 PMID: 30284484

Objective: To develop a nanoparticle-mediated drug delivery system for Tanshinone IIA.

Summary: Demonstrated a novel delivery mechanism of a therapeutic agent, tanshinone IIA-NP, which prevented IκB phosphorylation and subsequent NF-κB activation resulting in the suppression of inflammatory responses and reduction of cardiomyocytes apoptosis and fibrotic process, contributing to the mitigation of LV remodeling and improvement of cardiac functions following MI.

Usage: Immunoblotting analysis for the activation of AT-1R on an ischemic border zone 4 weeks after explosion to coronary artery ligation.

Related Products: Angiotensin II receptor (AT-1R) Rabbit Polyclonal, affinity-purified (Cat. #AB-N27AP)

Sabbah S, Papendorp C, Koplas E, Beltoja M, Etebari C, Gunesch AN, Carrete L, Kim MT, Manoff G, Bhatia-Lin A, Zhao T, Schreck D, Dowling H, Briggman KL, Berson DM (2018) Synaptic circuits for irradiance coding by intrinsically photosensitive retinal ganglion cells. bioRxiv 442954. doi: 10.1101/442954

Objective: To explore the synaptic networks responsible for the unique capacity of intrinsically photosensitive retinal ganglion cells (ipRGCs) to encode overall light intensity. This luminance signal is crucial for circadian, pupillary and related reflexive responses light.

Summary: Most ipRGCs sample from all bipolar terminals costratifying with their dendrites, but M1 cells avoid all OFF bipolar input and accept only ectopic ribbon synapses from ON cone bipolar axonal shafts. These monad synapses are equipped with as many as a dozen ribbons and only one postsynaptic process.

Usage: Immunohistochemistry of retina – after recording, retinas were fixed and counterstained with rabbit anti-melanopsin (1:1000) to enhance the fluorescence of the GFP-based GCaMP6f indicator.

Related Products: Melanopsin Rabbit Polyclonal (Cat. #AB-N38), Metabotropic Glutamate Receptor 2 (mGluR2) Mouse Monoclonal (Cat. #AB-N32)