-
Your selected country is
Singapore
- Change country/language
-
Reagents
- Flow Cytometry Reagents
-
Western Blotting and Molecular Reagents
- Immunoassay Reagents
-
Single-Cell Multiomics Reagents
- BD® AbSeq Assay
- BD Rhapsody™ Accessory Kits
- BD® Single-Cell Multiplexing Kit
- BD Rhapsody™ Targeted mRNA Kits
- BD Rhapsody™ TCR/BCR Profiling Assays for Human and Mouse
- BD Rhapsody™ Whole Transcriptome Analysis (WTA) Amplification Kit
- BD® OMICS-Guard Sample Preservation Buffer
- BD Rhapsody™ ATAC-Seq Assays
-
Functional Assays
-
Microscopy and Imaging Reagents
-
Cell Preparation and Separation Reagents
-
Training
- Flow Cytometry Basic Training
-
Product-Based Training
- FACSAria Product Based Training
- FACSMelody Product-Based Training
- FACSLyric Product-Based Training
- FACSCanto Product-Based Training
- LSRFortessa Product-Based Training
- FACSymphony Product-Based Training
- FACSDuet Product-Based Training
- HTS Product-Based Training
- BD FACSDiscover™ S8 Cell Sorter Product Training
-
Advanced Training
-
Thought Leadership
-
Product News
- Blogs
- Scientific Publications
-
Events
- Nature Research Academies Workshop 2023
- CYTO 2023: Advancing the World of Cytometry
- Singapore Gene & Cell Therapy Conference 2023
- EuroFlow Educational Workshop
- Nature Research Masterclass 2023
- Novel Approaches to Single-Cell Plant Research: from Real-Time Imaging Cell Sorting to Single-Nuclei Transcriptomics
- Advances in Immune Monitoring Series
-
Product News
-
- BD® AbSeq Assay
- BD Rhapsody™ Accessory Kits
- BD® Single-Cell Multiplexing Kit
- BD Rhapsody™ Targeted mRNA Kits
- BD Rhapsody™ TCR/BCR Profiling Assays for Human and Mouse
- BD Rhapsody™ Whole Transcriptome Analysis (WTA) Amplification Kit
- BD® OMICS-Guard Sample Preservation Buffer
- BD Rhapsody™ ATAC-Seq Assays
-
- FACSAria Product Based Training
- FACSMelody Product-Based Training
- FACSLyric Product-Based Training
- FACSCanto Product-Based Training
- LSRFortessa Product-Based Training
- FACSymphony Product-Based Training
- FACSDuet Product-Based Training
- HTS Product-Based Training
- BD FACSDiscover™ S8 Cell Sorter Product Training
-
- Nature Research Academies Workshop 2023
- CYTO 2023: Advancing the World of Cytometry
- Singapore Gene & Cell Therapy Conference 2023
- EuroFlow Educational Workshop
- Nature Research Masterclass 2023
- Novel Approaches to Single-Cell Plant Research: from Real-Time Imaging Cell Sorting to Single-Nuclei Transcriptomics
- Advances in Immune Monitoring Series
- Singapore (English)
- Change country/language
Old Browser
This page has been recently translated and is available in French now.
Looks like you're visiting us from {countryName}.
Would you like to stay on the current country site or be switched to your country?
Immunofluorescence
Immunofluorescence Staining of Human Cells by Lysed Whole Blood Method
- Add 100 µl of well-mixed anticoagulated whole blood to the bottom of a labeled tube. (We use EDTA as the anticoagulant.)
- Add the appropriate primary antibody to each tube. If using unlabeled antibody, a titration is suggested. Conjugated antibody should be used as directed, usually 5 or 20 µl per sample for the pre-diluted test size products from BD Biosciences. Since applications vary, each investigator should titrate reagents to obtain optimal results.
- Mix well, then incubate in the dark at room temperature for 20-30 minutes.
- Remove tubes from dark chamber and mix each tube well. Add 2 ml of lysing solution to each tube. Vortex each tube.
- Incubate at room temperature in the dark for 10-15 minutes.
- Centrifuge for 5 minutes at 1000 rpm (200 x g).
- Remove supernatant by aspiration. Then vortex and add 2 ml washing solution to each tube.
- Centrifuge for 5 minutes at 1000 rpm (200 x g).
- Remove supernatant by aspiration.
- If required, add appropriate second step antibody (at optimal concentration) to each tube and vortex gently (if second step antibody is not required, proceed to step 18).
- Incubate in the dark at room temperature for 20-30 minutes.
- Remove from the dark.
- Mix well, then add 2 ml washing buffer to each tube.
- Centrifuge for 5 minutes at 1000 rpm (200 x g).
- Remove supernatant by aspiration and vortex.
- If a third step is not required, proceed to step 18. For third step, add an appropriate third-step reagent (at optimal concentration) to each tube and vortex gently.
- Repeat steps 11-15.
- If you will analyze the same day, add 500 µl wash buffer to each tube, vortex, and analyze within 8 hours. If not, fix cells by resuspending in 2% paraformaldehyde buffer* (30 minutes, room temperature), and wash with wash buffer. Resuspend cells in 500 µl wash buffer and store in the refrigerator at 2-8°C for up to 36 hours.
*Extended storage of fluorescent dyes in paraformaldehyde may affect fluorescence. We recommend using our BD Stabilizing Fixative (cat. no. 338036) for preserving immunofluorescent staining (see the TDS for detailed instructions).
Solutions:
RBC Lysing Solution: BD PharmLyse (cat. no. 555899) or BD FACS Lysing Solution (cat. no. 349202).
Washing Solution: PBS + 0.1% sodium azide + 1% fetal bovine serum.
Paraformaldehyde buffer: 2% paraformaldehyde in PBS.
BD Stabilizing Fixative (cat. no. 338036).
Report a Site Issue
This form is intended to help us improve our website experience. For other support, please visit our Contact Us page.