In This Issue

FEATURED NEW PRODUCTS




Bring the Power of Fluorescence Quantitation to Your Bench — The Qubit® 2.0 Fluorometer


 
Reliable Cell Viability Data in Only 10 Minutes
— PrestoBlue™ Cell Viability Reagent

 
Simple, Portable Signal Transduction Assays — BacMam-Enabled Cellular Assays

 New Antibodies for Hyperosmotic Stress Research — ABfinity™ Recombinant Rabbit Monoclonal Antibodies



Brighter Apoptosis Detection — New Alexa Fluor® 488–Annexin V Apoptosis Detection Kits




Protein Sample Prep Gets a Premiere Upgrade — Click-iT® Protein Enrichment Kit



DEPARTMENTS



OTHER PUBLICATIONS


BioProbes 63  Bioprobes®
New! Expanded Content Now Online



The Molecular Probes® Handbook
11th Edition


FEATURED NEW PRODUCTS

AVAILABLE NOVEMBER 2010 — Bring the Power of Fluorescence Quantitation to Your Bench — The Qubit® 2.0 Fluorometer

what it is
The Qubit® 2.0 Fluorometer is the next generation in DNA, RNA, and protein quantitation. Central to the Qubit® Fluorometric Quantitation Platform, this instrument was designed with one thing in mind: to enable you to easily and accurately quantitate your nucleic acid or protein sample, to help avoid wasted samples or time repeating experiments.



what it offers

  • Accurate, specific, and sensitive—fluorescence-based dyes bind only to DNA, RNA, or protein
  • Easy to use—improved, intuitive touch-screen interface
  • Save your data—data logging function and USB port

how it works
The Qubit® 2.0 Fluorometer is an easy-to-use analytical instrument featuring advanced optics, data analysis algorithms, and an intuitive touch-screen user interface. The instrument is designed to work seamlessly with Qubit™ assays for DNA, RNA, and protein quantitation. The integrated design of the instrument and the assays makes the Qubit® 2.0 Fluorometer far more sensitive and accurate than UV absorbance; this helps to avoid the repetition of experiments due to inaccurate nucleic acid or protein measurements.

  • Notify me when the Qubit® 2.0 Fluorometer is available
Qubit 2.0

The Qubit® 2.0 Fluorometer.

Reliable Cell Viability Data in Only 10 Minutes — PrestoBlue™ Cell Viability Reagent

what it is
The PrestoBlue™ reagent is a resazurin-based solution that functions as a cell viability indicator. The reagent contains a cell-permeant compound that is blue and virtually nonfluorescent. When added to cells, the reagent is modified by the reducing environment of the viable cell, turns red, and becomes highly fluorescent.



what it offers

  • Save time—incubation step as short as 10 minutes
  • High-quality results—large assay window and z’ values
  • Experience convenience—homogeneous add-and-read format
how it works
The PrestoBlue™ reagent is a simple add-and-read cell viability assay. Once a plate containing cells and compounds is prepared, the PrestoBlue™ reagent is added, and results can be read in as little as 10 minutes. PrestoBlue™ reagent allows for reliable, high-quality cell viability data and continuous live-cell monitoring and assessment after measuring viability.

PrestoBlue™ Cell Viability Reagent

Reliable, high-quality cell viability data.
Jurkat cells were plated in cell culture medium in a 384-well plate in quadruplicate starting at 100,000 cells/well with a 2-fold serial dilution. Fluorescence measurements were taken after 10 min of incubation and again after 16 hr of incubation with PrestoBlue™ reagent.
Product
Quantity
Cat. No.
PrestoBlue™ Cell Viability Reagent
25 mL
A13261Order Now
PrestoBlue™ Cell Viability Reagent
100 mLA13262Order Now

Simple, Portable Signal Transduction Assays — BacMam-Enabled Cellular Assays

what they are
BacMam-enabled Cellular Assays are an efficient, easy-to-optimize, and robust method for interrogating specific signal transduction events in a cell background of choice. These assays combine BacMam-mediated gene delivery and LanthaScreen® TR-FRET technology to provide unique benefits including portability into different cell types and a quick, HTS-amenable assay readout.



what they offer

  • Portable—compatible with most cell types, including primary and stem cells
  • Fast—develop assays typically in less than 1 week, without having to generate stable cell lines
  • Flexible—measure multiple modifications such as phosphorylation, acetylation, or ubiquitination
how they work
Compared to other types of cellular assays, BacMam-enabled Cellular Assays reduce assay development time by minimizing the need to generate a stable cell line and allowing a choice of cellular background. Once cells are transduced with the BacMam reagent, LanthaScreen® technology is utilized for the readout. This simple, “addition-only” protocol requires minimal hands-on time.

BacMam-enabled Cellular Assay workflow

BacMam-enabled Cellular Assay workflow.
Cells are treated with BacMam reagent encoding a GFP-fusion protein and plated in 384-well format. 24-hours posttransduction, the cells are stimulated to induce the posttranslational modification of the GFP-substrate (e.g., phosphorylation, as shown). Cells are then lysed in the presence of a terbium anti–modification-specific antibody prior to the LanthaScreen® assay readout.
Product
Quantity
Cat. No.
BacMam Histone H3 [AcLys9] Cellular Assay Kit
1 kit
A12897Order Now
BacMam Histone H3 [pSer10] Cellular Assay Kit
1 kitA12898Order Now
BacMam AKT [pSer473] Cellular Assay Kit
1 kitA12899
Order Now
BacMam AKT [pThr308] Cellular Assay Kit
1 kitA12900
Order Now
BacMam p53 [AcLys382] Cellular Assay Kit
1 kitA12901
Order Now
BacMam p53 [pSer15] Cellular Assay Kit
1 kitA12902
Order Now

New Antibodies for Hyperosmotic Stress Research — ABfinity™ Recombinant Rabbit Monoclonal Antibodies

what it is
FAK phosphorylation at tyrosine 861 regulates hyperosmotic stress signaling and has been indicated as a requirement for mammalian cells to survive under hyperosmotic stress. EGFR is also activated in downstream hyperosmotic stress signaling cascades. Invitrogen now offers two new ABfinity™ rabbit recombinant monoclonal antibodies to help elucidate phosphor-FAK Y861 and EGFR signaling.



what it offers

  • Consistency—recombinant antibodies enable consistent results

how it works
ABfinity™ recombinant rabbit monoclonal antibodies help ensure consistent antibody performance lot after lot, so you don’t have to revalidate dilutions for your experiments when you order more. FAK [pY861] ABfinity™ Recombinant Rabbit Monoclonal Antibody is phosphospecific to tyrosine 861 and will not cross-react with the non-phosphorylated Y861 form of FAK. The EGFR and FAK [pY816] ABfinity™ antibodies are validated in western blotting and immunofluorescence applications.

Immunocytochemistry of HeLa cells

Immunocytochemistry of HeLa cells labeled with rabbit anti-EGFR.
HeLa cells were labeled with rabbit anti-EGFR (10 µg/mL). Alexa Fluor® 488 goat anti–rabbit IgG was used at 1:1,000 as the secondary antibody. Shown is a composite image of DAPI (blue), anti-EGFR (green), and Alexa Fluor® 594 Phalloidin (red).
Product
Quantity
Cat. No.
FAK [pY861] ABfinity™ Recombinant Rabbit Monoclonal Antibody
100 µg
700154Order Now
EGFR ABfinity™ Recombinant Rabbit Monoclonal Antibody
100 µg700308Order Now

Brighter Apoptosis Detection — New Alexa Fluor® 488–Annexin V Apoptosis Detection Kits

what they are
The new Alexa Fluor® 498–Annexin V Apoptosis Detection Kits detect apoptotic cells using recombinant annexin V conjugated to the Alexa Fluor® 488 dye, and dead cells using propidium iodide (PI). After treatment with both probes, apoptotic cells fluoresce green, dead cells fluoresce red and green, and live cells show little or no fluorescence.



what they offer

  • Better data—the use of Alexa Fluor® 488 dye produces brighter, more photostable Annexin V conjugates
  • Simple conversion—Alexa Fluor® 488 dye easily fits into the same filter configuration as FITC
  • Convenience—ready-to-use assays with validated flow cytometry protocols
  • More economical—reduced price per test for larger and smaller kit formats
how they work
In apoptotic cells, phosphatidyl serine (PS) is translocated from the inner to the outer leaflet of the plasma membrane, thus becoming exposed to the extracellular environment. Annexin V labeled with a fluorophore or biotin can be used to identify apoptotic cells by binding to extracellular PS. The Alexa Fluor® 488 dye has proven to make brighter and more photostable bioconjugates than fluorescein (FITC), providing better separation and population resolution. Propidium iodide stains necrotic cells with red fluorescence.

Apoptosis detection by flow cytometry

Apoptosis detection by flow cytometry. 
Camptothecin-induced Jurkat cells were stained with the recommended concentration of annexin V conjugate for 30 min, then washed, resuspended, and counterstained with propidium iodide. Cells were analyzed on the Attune™ Acoustic Focusing Cytometer using a 488 nm laser source and 530/30 nm and 575/24 nm emission filters.
Product
Quantity
Cat. No.
Dead Cell Apoptosis Kit with Annexin V Alexa Fluor® 488 and PI, 50 assays for flow cytometry
1 kit
V13241Order Now
Dead Cell Apoptosis Kit with Annexin V Alexa Fluor® 488 and PI, 250 assays for flow cytometry 
1 kitV13245Order Now
Annexin Binding Buffer, 5X concentrate for flow cytometry
1 kitV13246
Order Now

Protein Sample Prep Gets a Premiere Upgrade — Click-iT® Protein Enrichment Kit

what it is
The Click-iT® Protein Enrichment Kit allows for the efficient capture of click chemistry–based azide-modified proteins on a resin of alkyne agarose. Bound protein is then released by standard proteolytic release upstream of protein sequencing. Superior to biotin- or lectin-based approaches, the kit is ideal for proteomics, LC/MS/MS-based biomarker discovery, PTM analysis, and more.



what it offers

  • Allows for global differential profiling of newly expressed (nascent) and posttranslationally modified (PTM) proteins
  • Improves signal to noise by eliminating nonspecific interactions and increasing selectivity
  • Allows for characterization of protein and PTM stability and turnover rates
how it works
The azide click chemistry modification can occur via metabolic feeding, enzymatic addition, or chemical reaction. Click-azide modified proteins or their posttranslationally modified forms are enriched from protein extracts on the alkyne-agarose resin supplied with the kit. Once anchored to the resin via copper-catalyzed click chemistry, extensive washing yields a highly enriched population of nascent molecules that can be further characterized by mass spectrometry. The alkyne-agarose resin improves upon existing biotin approaches by >8 fold, with a signal-to-noise ratio of 25 for the Click-iT® resin vs. 3 for biotin.

Click-iT® Protein Enrichment Protocol Comparison

Comparison of Click-iT® protein enrichment technology and biotin-streptavidin protocols.

Product
Quantity
Cat. No.
Click-iT® Protein Enrichment Kit, for click chemistry capture of azide-modified proteins
1 kit
C10416Order Now

DEPARTMENTS

Buzzworthy


Recombinant phospholipase A1 (Ves v 1) from yellow jacket venom for improved diagnosis of hymenoptera venom hypersensitivity
Seismann H, Blank S, Cifuentes L et al. (2010) Clin Mol Allergy 8:7.

In certain individuals, the sting of a yellow jacket can induce life-threatening anaphylaxis. Efforts to develop reliable tests to identify sensitive individuals and produce efficient treatments for patients with yellow jacket stings have been hampered by the scarcity of purified marker allergens. In yellow jacket venom (YJV), phospholipase A1 (Ves v 1) and antigen 5 (Ves v 5) are primarily responsible for IgE-mediated allergic reactions, and these nonglycosylated proteins are unique candidates for diagnosis of YJV allergy. Although expression of Ves v 5 had previously been demonstrated, Ves v 1 could only be produced in an insoluble form. In a recent publication by Seismann and colleagues, the researchers were able to demonstrate the secretion of recombinant Ves v 1 and Ves v 5 in insect cells in soluble form. Using a colorimetric phosphlipase test (EnzChek® Phospholipase A1 Assay Kit, Cat. No. E10219/E10221) they showed that rVes v 1 retained its enzymatic activity, and both proteins were able to stimulate basophils isolated from venom-allergic patients. The authors concluded that purified recombinant allergens Ves v 1 and Ves v 5 hold promise as valuable tools in the search for yellow jacket venom allergy diagnosis and treatment.



Product
Quantity Cat. No.
EnzChek® Phospholipase A1 Assay kit for 2 plates
1 kit
E10219Order Now
EnzChek® Phospholipase A1 Assay kit for 10 plates
1 kit
E10221Order Now


Live-Cell Imaging
 Live-cell imaging of neurons using CellLight® MAP4-RFP. Hippocampal tissue from postnatal day 3 rats was harvested and dissociated in neural culture medium, then plated onto glial feeder cultures. Cells were resuspended at 50,000 cells/mL in complete neural culture medium plus mitotic inhibitors. The medium was removed from the glial feeder cultures and replaced with 2 mL of the neural cell suspension. CellLight® MAP4-RFP (50 μL) was added to the plates, and cells were incubated overnight. Fluorescence was observed in neurons after 48 hr in culture.

Product
Quantity
Cat. No.
CellLight® MAP4-RFP, BacMam 2.0
1 mL
C10599Order Now

Attune™ Acoustic Cytometer

Join the Attune™ Cytometer Revolution
The positive reviews are rolling in from researchers who’ve experienced the Attune™ Acoustic Focusing Cytometer.

“Overall, I would say that the Attune seems to be not only revolutionary, but…also a significant step in the improvement of the existing technology.”—Tomas Adejumo, UCL Scientific Support Services, posted on the GeneRoom.net website.

During a recent installation, a researcher was so anxious to get started on the new technology he remarked to the installing engineer: “You make it work, I will make it famous!” Other customers are excited about the blue/violet laser configuration, which enables them to monitor cell health, proliferation, apoptosis, and cell cycle, along with their fluorescent proteins excited by the blue laser. Stay tuned for publications and more comments from thrilled Attune™ Cytometer customers.




Flow Cytometry Catalog
New Resource for Cell Cycle Analysis by Flow Cytometry

Flow cytometry provides a powerful tool to assess cells in G0/G1 phase versus the S phase, the G2 phase, or polyploidy. Now we’ve made it easier to select the right cell cycle assay for your flow cytometry experiments:

  • Live cell cycle assays
  • Fixed cell cycle assays
  • Antibodies for cell cycle markers


ADME/Tox News


Molecular Probes® The Handbook

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