Recombinant Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [28-8] to PD-L1 - BSA and Azide free
- Suitable for: WB, ICC/IF, Flow Cyt, IHC-P
- Knockout validated
- Reacts with: Human
Related conjugates and formulations
Overview
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Product name
Anti-PD-L1 antibody [28-8] - BSA and Azide free
See all PD-L1 primary antibodies -
Description
Rabbit monoclonal [28-8] to PD-L1 - BSA and Azide free -
Host species
Rabbit -
Tested applications
Suitable for: WB, ICC/IF, Flow Cyt, IHC-Pmore details -
Species reactivity
Reacts with: Human -
Immunogen
Recombinant full length protein. This information is proprietary to Abcam and/or its suppliers.
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Positive control
- Tissue: Human tonsil and head and neck squamous cell carcinoma tissues; L2987 cell line. Cell Lines: Positives: B-CPAP- high, ES-2- medium, HCC70 - low CHO-PDL1 For additional information, please refer to here: Programmed death-ligand 1 (PD-L1) expression in various tumor types - http://www.immunotherapyofcancer.org/content/1/S1/P53
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General notes
ab228413 is the carrier-free version of abab205921.
Additional information on positive controls:
Tissue:
Tonsil- with hyperreactive changes
Note: Tonsil Specimens- is recommended to screen several hyper-reactive tonsils to find those with highest expression of PD-L1 in crypt epithelium, macrophages homing the germinal centers and interfollicular mononuclear leukocytes.
Tumor tissues- prescreened for positive tumor and inflammatory infiltrates
Note: Tumor Specimens- PD-L1 expression varies by tumor type so screening is recommended to find positive and negative tumor controls. Refer to web link publication below to find some suggested tumor types. Many tumor specimens have some inflammatory macrophages and mononuclear leukocytes. Look for specimens with high numbers of these cells
Cell Lines: Positives: B-CPAP- high, ES-2- medium, HCC70 - lowFor IHC on FFPE tissues, antigen retrieval buffer (ab208572) and IHC detection kit HRP/DAB (ab209101) is recommended.
For primary negative control, isotype control, RabMAb negative control antibody (ab172730) is recommended.
For negative control sample, use cell line COLO205, see ab95363.
For PD-L1 protein, see ab167713Recommended protocols:
For IHC usage on FFPE tissues, the following antigen solution is recommended with clone 28-8 - Universal HIER antigen retrieval reagent (ab208572)Western blot usage
For clone 28-8, it is recommended to use Odyssey system. This system has the advantages of a wider dynamic range and less background than chemiluminescence.Anti-PD-L1 antibody [28-8] has been used as detector antibody in Human PD-L1 SimpleStep ELISA® kit (ab214565).
Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.
This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.
Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.
This product is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar® is a trademark of Fluidigm Canada Inc.
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C. Do Not Freeze. -
Storage buffer
pH: 7.20
Constituent: 100% PBS -
Carrier free
Yes -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
28-8 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
- Anti-PD-L1 antibody [28-8] (ab205921)
- APC Anti-PD-L1 antibody [28-8] (ab206967)
- Anti-PD-L1 antibody [28-8] - Low endotoxin, Azide free (ab209889)
- Alexa Fluor® 488 Anti-PD-L1 antibody [28-8] (ab209959)
- Alexa Fluor® 647 Anti-PD-L1 antibody [28-8] - Extracellular domain (ab209960)
- HRP Anti-PD-L1 antibody [28-8] (ab209961)
- PE Anti-PD-L1 antibody [28-8] (ab209962)
- Alexa Fluor® 555 Anti-PD-L1 antibody [28-8] (ab213358)
- Alexa Fluor® 568 Anti-PD-L1 antibody [28-8] (ab213359)
- Alexa Fluor® 594 Anti-PD-L1 antibody [28-8] (ab213360)
- FITC Anti-PD-L1 antibody [28-8] (ab224027)
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Compatible Secondaries
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Conjugation kits
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Immunohistochemistry kits
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Isotype control
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Recombinant Protein
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Related Products
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Unmasking reagent
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab228413 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application | Abreviews | Notes |
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WB |
Use at an assay dependent concentration. Predicted molecular weight: 33 kDa.
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ICC/IF |
Use at an assay dependent concentration.
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Flow Cyt |
Use at an assay dependent concentration.
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
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IHC-P | (1) |
Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
For antigen buffer for FFPE tissue, it is recommended to use Universal HIER antigen retrieval reagent (ab208572). |
Notes |
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WB
Use at an assay dependent concentration. Predicted molecular weight: 33 kDa. |
ICC/IF
Use at an assay dependent concentration. |
Flow Cyt
Use at an assay dependent concentration. ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. For antigen buffer for FFPE tissue, it is recommended to use Universal HIER antigen retrieval reagent (ab208572). |
Target
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Function
Involved in the costimulatory signal, essential for T-cell proliferation and production of IL10 and IFNG, in an IL2-dependent and a PDCD1-independent manner. Interaction with PDCD1 inhibits T-cell proliferation and cytokine production. -
Tissue specificity
Highly expressed in the heart, skeletal muscle, placenta and lung. Weakly expressed in the thymus, spleen, kidney and liver. Expressed on activated T- and B-cells, dendritic cells, keratinocytes and monocytes. -
Sequence similarities
Belongs to the immunoglobulin superfamily. BTN/MOG family.
Contains 1 Ig-like C2-type (immunoglobulin-like) domain.
Contains 1 Ig-like V-type (immunoglobulin-like) domain. -
Cellular localization
Cell membrane and Endomembrane system. - Information by UniProt
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Database links
- Entrez Gene: 29126 Human
- Omim: 605402 Human
- SwissProt: Q9NZQ7 Human
- Unigene: 521989 Human
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Alternative names
- B7 H antibody
- B7 H1 antibody
- B7 homolog 1 antibody
see all
Images
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
Immunohistochemical analysis of formalin-fixed, paraffin-embedded L2987 (Human lung adenocarcinoma cell line with endogenous PD-L1 expression) cells labeling PD-L1 with ab205921 at 2 µg/ml. Counterstained with Hematoxylin.
For antigen retrival buffer, Universal HIER antigen retrieval reagent (ab208572) was used.
For IHC detection kit, Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) is recommended.This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
Immunohistochemical analysis of formalin-fixed, paraffin-embedded Human head and neck squamous cell carcinoma tissue labeling PD-L1 with ab205921 at 2 µg/ml. Counterstained with Hematoxylin.
For antigen retrival buffer, Universal HIER antigen retrieval reagent (ab208572) was used.
For IHC detection kit, Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) is recommended.This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human melanoma tissue labelling PD-L1 with ab205921 on Ventana Ultra. Tumor cells and immune cells show PD-L1 positive plasma membrane staining.
For antigen retrival buffer, Universal HIER antigen retrieval reagent (ab208572) was used.
For IHC detection kit, Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) is recommended.This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human non-small cell lung cancer tissue labelling PD-L1 with ab205921. Staining can be seen in tumor associated macrophages and tumor cells.
For antigen retrival buffer, Universal HIER antigen retrieval reagent (ab208572) was used.
For IHC detection kit, Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) is recommended.This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
Immunohistochemical analysis of Human Lung NSCLC with ab205921 at 2 µg/ml.
High power view
A) Rabbit IgG, 5 µg/mL. No staining
B) Anti PD-L1, 2 µg/mL (ab205921 batches 1)
C) Anti PD-L1, 2 µg/mL (ab205921 batches 3)
D) Anti PD-L1, 2 µg/mL (ab205921 batches 4)
E) Anti PD-L1, 2 µg/mL (ab205921 batches 5)
F) Anti PD-L1, 2 µg/mL (ab205921 batches 6)All batches/lots (1,3,4,5,6) showed consistent results.
Note linear and complete or partial (arrows) PD-L1 staining of tumor cells. Tumor associated immune cells localized over the tumor margin exhibit positive plasma membrane staining (small arrows).For recommended Immunohistochemistry (IHC) protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy).
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
Immunohistochemical analysis of CHO Parental cells with ab205921 at 2 µg/ml.
High power view
A) Rabbit IgG, 5 µg/mL. No staining
B) Anti PD-L1, 2 µg/mL (ab205921 batches 1)
C) Anti PD-L1, 2 µg/mL (ab205921 batches 3)
D) Anti PD-L1, 2 µg/mL (ab205921 batches 4)
E) Anti PD-L1, 2 µg/mL (ab205921 batches 5)
F) Anti PD-L1, 2 µg/mL (ab205921 batches 6)
G) Anti PD-L1, 2 µg/mL (ab205921 batches 7)All batches/lots (1,3,4,5,6,7) showed consistent results.
Note absence of PD-L1 expression in CHO parental cells.
For recommended Immunohistochemistry (IHC) protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy).
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
Immunohistochemical analysis of CHO PD-L1 cells with ab205921 at 2 µg/ml.
High power view
A) Rabbit IgG, 5 µg/mL. No staining
B) Anti PD-L1, 2 µg/mL (ab205921 batches 1)
C) Anti PD-L1, 2 µg/mL (ab205921 batches 3)
D) Anti PD-L1, 2 µg/mL (ab205921 batches 4)
E) Anti PD-L1, 2 µg/mL (ab205921 batches 5)
F) Anti PD-L1, 2 µg/mL (ab205921 batches 6)
G) Anti PD-L1, 2 µg/mL (ab205921 batches 7)All batches/lots (1,3,4,5,6,7) showed consistent results.
Note strong, moderate, and weak (red, yellow, and white arrows respectively) plasma membrane staining of CHO PD-L1 transfected cellsFor recommended Immunohistochemistry (IHC) protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy).
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
Immunohistochemical staining of PD-L1 in formalin fixed, paraffin embedded human non-squamous non-small cell lung cancer (NSQ-NSCLC) using ab205921 at a dilution of 1/400, incubated for an hour at room temperature. Heat mediated antigen retrieval was carried out in low pH buffer and the sample was blocked with peroxidase blocking buffer for 3 minutes.
This image was courteously provided by Dr. Kai Schmitt from the Institute of Pathology, Saarbrücken-Rastpfuhl.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
Ab205921 specificity testing by Immunohistochemistry (KO testing): Loss of detection on KO Cells
Strong IHC detection with anti-PD-L1 (ab205921, clone 28-8) is seen in human lung adenocarcinoma tumor cell line L2987. PDL1 gene was edited in L2987 cells using TALEN constructs targeting exon4 of human PD-L1, transcript variant 1 (NM_014143.3) and complete knock out (K.O) confirmed by deep sequencing in clone L2-14. IHC detection is completely eliminated in the L2987 L2-14 K.O. cell line.For recommended Immunohistochemistry (IHC) protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy).
Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
Anti-PD-L1 antibody [28-8] (ab205921)
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human tonsil tissue labelling PD-L1 with ab205921 at a dilution of 1:400. Heat mediated antigen retrieval was performed using AR9 antigen retrieval solution, and microwave treatment for 15 min at 20% power. Anti-Rabbit/Mouse HRP polymer (PerkinElmer Opal Polymer HRP Ms Plus Rb) was used as secondary antibody. Opal tyramide amplification was performed using Opal 520 fluorophore. Counterstained with DAPI stain. Image scanned with Vectra 3.0 and analyzed via Phenochart software.
This image was courteously provided by Dr. Houssein Abdul Sater, Georgia Cancer Center.This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
IHC image of ab205921 staining PD-L1 in PD-L1 Dynamic Range Analyte Control formalin fixed paraffin embedded cell lines (HistoCyte Laboratories), performed on a Leica BOND RX (Polymer Refine kit). The section was pre-treated using heat mediated antigen retrieval with EDTA buffer (pH9, epitope retrieval solution 2) for 30 mins at 98°C. The section was then incubated with ab205921, 5μg/ml working concentration, for 60 mins at room temperature and detected using an HRP conjugated compact polymer system for 8 minutes at room temperature. DAB was used as the chromogen for 10 minutes at room temperature. The section was then counterstained with hematoxylin, blued, dehydrated, cleared and mounted with DPX.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
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ab205921 specificity testing by Flow Cytometry (KO testing): Loss of detection on KO cells.
Strong detection with anti-PD-L1 (ab205921, clone 28-8) TALEN constructs targeting exon4 of human PD-L1, transcript variant 1 (NM_014143.3) and complete knock out (K.O) confirmed by deep sequencing in clone L2-14. Cell surface staining is almost completely eliminated in the L2987 L2-14 KO cell line.For recommended Flow Cytometry (Flow Cyt) protocol, please refer to the protocol book in the protocol section and/or here (downloadable copy).
Alexa Fluor® 488 (ab209959) and Alexa Fluor® 647 (ab209960) conjugated versions are available for this clone.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [28-8] - BSA and Azide free (ab228413)
IHC image of ab205921 staining PD-L1 in human tonsil formalin fixed paraffin embedded tissue sections*, performed on a Leica BOND RX (Polymer Refine kit). The section was pre-treated using heat mediated antigen retrieval with EDTA buffer (pH9, epitope retrieval solution 2) for 30 mins at 98°C. The section was then incubated with ab205921, 5μg/ml working concentration, for 60 mins at room temperature and detected using an HRP conjugated compact polymer system for 8 minutes at room temperature. DAB was used as the chromogen for 10 minutes at room temperature. The section was then counterstained with hematoxylin, blued, dehydrated, cleared and mounted with DPX.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab205921).
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
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Datasheet download
Certificate of Compliance
References (0)
ab228413 has not yet been referenced specifically in any publications.