Intracellular ROS Assay

Reactive oxygen species (ROS) are active chemicals containing oxygen. They include peroxides, superoxides, hydroxyl radicals, singlet oxygen and α-Oxygen.

Creative Bioarray uses fluorescent probe DCFH-DA to detect reactive oxygen species. DCFH-DA itself has no fluorescence and can freely pass through the cell membrane. After entering the cell, it can be hydrolyzed by esterase in the cell to produce DCFH. DCFH can not penetrate the cell membrane, so the probe can be easily loaded into the cell. Reactive oxygen species in cells can oxidize non fluorescent DCFH to produce fluorescent DCF. The level of reactive oxygen species in cells can be known by detecting the fluorescence of DCF.

Service Process

Fig. 1 Trafficking of the extracellular and intracellular ROS and NO (Chen et al., 2017)

Step 1. Prepare single cell suspension (tissue sample)
Step 2. Hemolysis (this step is not necessary for cell samples)
Step 3. Centrifuge to remove supernatant
Step4. Adjust cell density
Step 5. Add the corresponding fluorescent probe
Step 6. Incubation
Step 7. Centrifuge the supernatant after washing
Step8. On machine detection

Provided by Customers

Instructions for specimen collection and preservation

In order to achieve better detection effect, please refer to the following principles to provide the collection and preservation of samples.

Tissue sample: fresh tissue shall be stored in liquid nitrogen or - 70 ℃, and the tissue shall not be less than 100mg (about the size of mung bean).
Cultured cells: take at least 2000000 cells in EP tube by cell counting, add 0.5ml normal saline or protein protective agent, mix well and store in refrigerator (- 70 ℃) to avoid repeated freezing and thawing.
Blood cell sample: save the blood sample with anticoagulant tube or separate the cells with lymphocyte separation solution, add 0.5ml normal saline or protein protective agent, and keep it (- 70 ℃ can be stored for a long time to avoid repeated freezing and thawing).
Serum or cell culture medium samples: after centrifugation to remove impurities, take the supernatant into the EP tube and store it (the above 4 ℃ can be stored for 15-20 days, and - 70 ℃ can be stored for a long time to avoid repeated freezing and thawing).

Instructions for sample transportation

In order to achieve better testing results, customers can send samples in any of the following ways.

Tissue samples and cell samples are transported with dry ice or with ice bags after adding protein protective agent.
Cell samples can also be sent directly to the culture bottle by express mail (sealed to ensure no pollution, no leakage of culture medium, cells do not grow too full, about 50%, filled with culture medium).
The serum or supernatant is sent directly in an ice bag (sealed to ensure that the liquid does not leak out, which can be reached in 2-3 days depending on the distance).

Inspection and Delivery Standards

Sample Type	Requirement	Transportation Conditions
Tissue	The isolated tissues were made into single cell suspension and added into PBS medium	Normal temperature transportation
Living cell	Sample size: 1 X 10⁶ cells / test, added into PBS medium	Normal temperature transportation
Blood	The blood was collected in the anticoagulant blood collection vessel of EDTA heparin sodium	Normal temperature transportation

Creative Bioarray is dedicated to providing high-quality products, comprehensive services, and tailored solutions to support and facilitate life sciences and pharmaceutical research and development. If you have any questions or needs, please contact us or make online inquiry, and our customer service staff will help you at the first time.

Reference

Chen, Y., Liu, H., Luo, X., Zhao, Z., Zou, Z., & Li, J. et al. (2017). The roles of reactive oxygen species (ROS) and autophagy in the survival and death of leukemia cells. Critical Reviews In Oncology/Hematology, 112, 21-30.

All services and products are for scientific use only, not for medical use！