Protein Transport Inhibitor Cocktail (500×) - sterile-filtered,BioReagent,500×

Cat. No.: P1508367

GRADE & PURITY Sterile-filtered, BioReagent, 500×

Synonyms

Flow Cytometry Protein Transport Inhibitor(Brefeldin A/Monensin Mixture) | Protein Transport Inhibitor Cocktail

Storage

Protected from light,Store at -20°C

Shipped In

Ice chest + Ice pads

Application

Flow Cytometry

Overview

Product Introduction

This product is a ready-to-use Protein Transport Inhibitor Cocktail (500X concentrate), containing optimized concentrations of Brefeldin A and Monensin Sodium. Working synergistically, they efficiently and specifically block the Golgi-mediated protein transport pathway within cells. It is widely used in intracellular detection assays for secretory proteins like cytokines and chemokines. When used in combination with cell stimulators (e.g., PMA/Ionomycin), it causes newly synthesized target proteins to accumulate intracellularly, making it a key reagent for Intracellular Cytokine Staining (ICS) by flow cytometry.

Key Components:

• Ethonal
• brefeldin A 1.5mg/mL，Monensin sodium salt 0.7mg/mL

Applications

• Co-stimulation Required: This is an inhibitor cocktail and must be used together with cell stimulators to be meaningful. Using it alone only blocks baseline secretion.
• Timing is Critical: Incubation time is key to success. Too short (<2 hours) leads to insufficient accumulation; too long (>6 hours) may increase non-specific background staining and affect viability.
• Supernatant Analysis: The secreted proteins blocked by this cocktail CANNOT be detected in the cell culture supernatant. Do not add this cocktail if you need to analyze factors secreted into the medium.
• Suitable for: Intracellular staining of both suspension cells (e.g., lymphocytes) and adherent cells.

Instructions for Use

Standard Protocol

• Cell Stimulation: Add stimulants (e.g., antigens, PMA/Ionomycin cocktail) to the cell culture to activate the cells.
• Add Inhibitors: At the beginning of stimulation (or shortly after), add this cocktail to the culture at a 1:500 dilution. For example, add 2 µL of the 500X concentrate to 1 mL of cell suspension.
• Incubation: Continue incubating the cells in a 37°C, 5% CO₂ incubator. Critical Incubation Time: Typically 4 to 6 hours. This allows sufficient protein synthesis and intracellular accumulation while minimizing the impact of inhibitor toxicity on cell viability. Incubation beyond 12 hours is not recommended.
• Post-Processing: After incubation, harvest the cells and proceed with standard protocols for surface staining, fixation, permeabilization, and intracellular staining.

Key Precautions

• Co-stimulation Required: This is an inhibitor cocktail and must be used together with cell stimulators to be meaningful. Using it alone only blocks baseline secretion.
• Timing is Critical: Incubation time is key to success. Too short (<2 hours) leads to insufficient accumulation; too long (>6 hours) may increase non-specific background staining and affect viability.
• Supernatant Analysis: The secreted proteins blocked by this cocktail CANNOT be detected in the cell culture supernatant. Do not add this cocktail if you need to analyze factors secreted into the medium.

• Aliquot upon receipt to minimize freeze-thaw cycles.

Specifications

Synonyms

Flow Cytometry Protein Transport Inhibitor(Brefeldin A/Monensin Mixture) | Protein Transport Inhibitor Cocktail

Specifications & Purity

sterile-filtered, BioReagent, 500×

Stability And Storage

Aliquot upon receipt and store frozen at -20°C(12 months). Protect from light. Avoid repeated freeze-thaw cycles.

Storage

Protected from light, Store at -20°C

Shipped In

Ice chest + Ice pads

This product requires cold chain shipping. Ground and other economy services are not available.

Grade

BioReagent, Sterile-filtered