Overview
Specifications
| Reagent | PULSin® |
|---|---|
| Molecule delivered | Protein; Antibody; Peptide |
| Cell types | Adherent & suspension cells grown in presence of serum |
| Number of transfection | 0.4 ml of PULSin® delivery reagent is sufficient to perform 25 delivery experiments in 6-well plates and 100 experiments in 24-well plates |
| Storage | Store PULSin® at 5 °C ± 3°C. Do not freeze. |
| Provided with | Hepes Buffer (20mM) for protein dilution; R-Phycoerythrin used as a positive control |
Summary
The delivery of protein and antibody using PULSin® represents a powerful approach for functional studies. For example, PULSin® enables you to study lethal proteins by controlling the level and time course of protein delivery into the cells. Similarly, delivery of blocking antibodies may provide additional information to traditional RNA interference experiments*. With PULSin® you can target intracellular proteins with antibodies in living cells without fixation step.
* G. Cassinelli et al. (2006). Inhibition of c-Met and prevention of spontaneous metastatic spreading by the 2-indolinone RPI-1, Mol Cancer Ther 5:9, 2388-2397.
Ordering information
| Reference number | Amount of reagent | Positive control (R-phycoerythrin) | Hepes dilution buffer |
|---|---|---|---|
| 101000010 | PULSin® 0.4 mL | 20 µg | 20 ml |
Description
Efficient delivery of proteins/antibodies and peptides to the cytoplasm
Proteins
PULSin® is able to deliver R-phycoerythrin, a fluorescent protein (240 kD), to the cytoplasm of up to 98% cells. As shown in Figure 1, the protein is evenly distributed in the cytoplasm and excluded from the nucleus due to its large size.
Fig 1: PULSin®-mediated intracellular delivery of R-phycoerythrin to NIH-3T3 cells. R-phycoerythrin (1 µg) is complexed with 4 µl of PULSin® for 15 min and added to NIH-3T3 cells in a 24-well plate. Live cells are observed by fluorescence microscopy after 16 h.
Antibodies
Antibodies are also successfully delivered to HeLa cells and able to recognize their target protein inside the cytoplasm.
For example, PULSin® permits the delivery of FITC-labeled anti-alpha-tubulin to the cytoplasm of 85% HeLa cells (Fig.2). Similarly, anti-giantin Alexa Fluor® 488 is delivered to the cytoplasm of 98% of live HeLa cells, labeling the Golgi apparatus (Fig.3).
|
Fig 2: Delivery of a fluorescein-conjugated anti-alpha-tubulin antibody with PULSin® to HeLa cells. |
Fig 3: Golgi labeling (green) of HeLa cells 24 h after delivery of 1 µg Alexa Fluor® 488 anti-Giantin using PULSin®. |
Plasma membrane is stained with ConA-rhodamine. Cells are observed by confocal microscopy.
Peptides
Peptides are biomolecules acting with high specificity at low concentrations. The delivery of substrate, inhibitor, modulator, or blocking peptides into cell allows protein function studies as well as the development of therapeutic approaches. PULSin® is able to successfully deliver Streptococcus TPE B epitope into HeLa cells (Fig. 4).
Fig 4: Delivery of Pep-A (Streptococcus TPE B epitope, 16 aa), into HeLa cells. Complexes are formed with Pep-A (1 µg, lissamine-rhodamine derivative, Sigma) and PULSin® (4 µl). Observation is carried out 16 h post-delivery.
Other proteins, antibodies and peptides can also be delivered to cells using PULSin® (Table 1).
Table 1: Examples of proteins, antibodies and peptides delivered to cells using PULSin®
PULSin® is easy to use and fast
PULSin® will save you time and efforts compared to other techniques using viral transduction or chemical conjugation. PULSin® reagent is ready-to-use and provided with a dilution buffer and a fluorescent control protein (R-phycoerythrin).
The protocol is fast: simply add the reagent to the protein, incubate and add to the cells.
Cells can be analyzed starting 4 hours post-delivery.
Fig 5: PULSin® – Protocol
Delivery to a wide variety of cells
PULSin® is able to deliver proteins and antibodies to a large variety of cell lines and primary cells (Table 2, Fig. 1-3).
Table 2: Efficiency of R-phycoerythrin delivery using PULSin® in selected cells.
Highly efficient transfer
The comparison of PULSin® with two other protein delivery reagents shows a higher efficiency of protein delivery (Fig. 5). Moreover, the amount of protein delivered per cell is higher with PULSin® as measured for R-phycoerythrin protein and for FITC-alpha-tubulin antibody (Fig. 6).
Fig 6: Comparison of PULSin® efficiency with two other protein delivery reagents. R-phycoerythrin (1 µg) is complexed with each reagent according to the manufacturer’s protocol. Complexes are added to HeLa cells and observed by fluorescence microscopy over 24 hours.
Fig 7: FACS analysis of HeLa cells after delivery of R-phycoerythrin (upper graph) or FITC-anti-alpha-tubulin antibody (lower graph) with PULSin® or with competitor reagents B and C. Data are presented as histograms of the mean intensity of fluorescence for each cell population.
Mechanism of delivery
PULSin® contains a proprietary cationic amphiphile molecule that forms non-covalent complexes with proteins and antibodies.
Complexes are internalized via anionic cell-adhesion receptors and are released into the cytoplasm where they disassemble. The process is non-toxic and delivers functional proteins.
FAQ
If you have any questions regarding PULSin®, please visit our dedicated Frequently asked questions or contact us.
Applications
Protein delivery
The intracellular delivery of proteins to live cells offers a powerful alternative as a scientific research tool where other approaches (DNA transfection, RNAi transfection) have failed.
Antibody delivery
The ability to introduce antibodies to live cells opens new insights to a wide range of applications such as protein intracellular trafficking studies, protein interference studies with blocking antibodies, live immunolabelling or protein phosphorylation states studies.
📰 Read Application note: Improving expression and purification of mAb using FectoCHO® Expression System and MabXpure™.
Quality
Every batch of PULSin® is tested by delivering R-phycoerythrin into HeLa cells.
PULSin® is provided as an aqueous solution in sterile and apyrogenic water. PULSin®, R-phycoerythrin and Hepes buffer are shipped at 4°C, should be stored at 4°C upon arrival, and as guaranteed by the Certificate of Analysis, will be stable for at least one year when stored appropriately.
Polyplus-transfection® has been awarded ISO 9001 Quality Management System Certification since 2002, which ensures that the company has established reliable and effective processes for manufacturing, quality control, distribution and customer support.
Protocol
In order to download a product protocol or a certificate of analysis, please create an account on Polyplus ® Portal .
Why would you need to create an account?
In this personal area you will have access to:
- Product Protocols
- Certificates of Analysis
- Exclusive webinars/articles
- And surprise features!
Other files
Related blog posts
Bibliography
| Cell Line | in vitro in vivo | Delivered Molecule | Reagent | Results & Citations | |
|---|---|---|---|---|---|
| EA.hy926, THP-1 | in vitro | Protein/Peptide/Antibody | PULSin | Bradburne, C. E. et al. (2013) Bioconjug Chem 24, 1570-83 Cytotoxicity of quantum dots used for in vitro cellular labeling: role of QD surface ligand, delivery modality, cell type, and direct comparison to organic fluorophores | More details |
| HC, Huh7, HuS-E/2 | in vitro | Protein/Peptide/Antibody | PULSin | Fukuhara, T. et al. (2011) Microbes Infect 13, 405-12 Intracellular delivery of serum-derived hepatitis C virus | More details |
| HEK-293 | in vitro | Protein/Peptide/Antibody | PULSin | Ohnuma, T. et al. (2010) Toxicol Appl Pharmacol 244, 27-36 Activation of the Nrf2/ARE pathway via S-alkylation of cysteine 151 in the chemopreventive agent-sensor Keap1 protein by falcarindiol, a conjugated diacetylene compound | More details |
| HtTA | in vitro | Protein/Peptide/Antibody | PULSin | Wang, C. H. et al. (2014) J Biomed Sci 21, 36 Phospholipase A/Acyltransferase enzyme activity of H-rev107 inhibits the H-RAS signaling pathway | More details |
| N2A, NSC-34 | in vitro | Protein/Peptide/Antibody | PULSin | Cascella, R. et al. (2017) FASEB J 31, 5609-5624 Quantitative assessment of the degradation of aggregated TDP-43 mediated by the ubiquitin proteasome system and macroautophagy | More details |
| RLE-6TN | in vitro | Protein/Peptide/Antibody | PULSin | Mazzocchi, L. C. et al. (2017) Am J Physiol Lung Cell Mol Physiol 313, L807-L824 TGF-beta inhibits alveolar protein transport by promoting shedding, regulated intramembrane proteolysis, and transcriptional downregulation of megalin | More details |
| Human microvascular endothelial cells | in vitro | Protein/Peptide/Antibody | PULSin | Thuringer, D. et al. (2015) Oncotarget , Primary tumor- and metastasis-derived colon cancer cells differently modulate connexin expression and function in human capillary endothelial cells | More details |
| Crayfish hematopoietic cells | in vitro | Protein/Peptide/Antibody | PULSin | Li, Y. Y. et al. (2019) Dev Comp Immunol 91, 101-107 Identification of a CqCaspase gene with antiviral activity from red claw crayfish Cherax quadricarinatus | More details |
| HEK-293 | in vitro | Protein/Peptide/Antibody | PULSin | Lingel A. et al. (2020) J Biol Chem , Amyloid precursor protein is a restriction factor that protects against Zika virus infection in mammalian brains | More details |
| Crustacean Hpt cell | in vitro | Protein/Peptide/Antibody | PULSin | Zheng SC. et al. (2019) Fish Shellfish Immunol 94, 934-943 A novel RING finger protein CqRNF152-like with self-ubiquitination activity inhibits white spot syndrome virus infection in a crustacean Cherax quadricarinatus | More details |