Anti-human IFN-α mAb (MT2/4/6), biotin

Anti-human IFN-α mAb (MT2/4/6), biotin

3425-6-250

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Content

Monoclonal antibody MT2/4/6, biotinylated. Supplied at 1 mg/ml in PBS with 0.02% sodium azide.

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Product specifications

Intended use

These monoclonal antibodies enable specific detection of human IFN-α subtypes 1/13, 2, 4, 5, 6, 7, 8, 10, 14, 16 and 17 (pan-specific detection) in immunoassays such as ELISpot, FluoroSpot, and ELISA.

Serum/Plasma samples

Recommendation

MT2/4/6 are recommended as detection mAb in ELISpot, FluoroSpot, and ELISA in combination with capture mAb MT1/3/5 (product code 3425-3).

Product details

ProductAnti-human IFN-α mAb (MT2/4/6), biotin
ApplicationELISpot, FluoroSpot, ELISA
AnalyteIFN-α pan
AntibodyMT2/4/6
ConjugateBiotin
ClonalityMonoclonal
ImmunogenRecombinant human IFN-α
HostMouse
IsotypeIgG1
ReactivityHuman, Non-human primates
Specificity

Native and recombinant human IFN-α. MT2/4/6 in combination with antibodies MT1/3/5 binds subtypes 1/13, 2, 4, 5, 6, 7, 8, 10, 14, 16 and 17.

Cross-reactivity

Cross-reacts with native IFN-α from rhesus macaque and cynomolgus macaque. For information on reactivity with other non-human primates (NHP), please check the NHP cross-reactivity guide.

Purification

Purified from in vitro cultures by protein G affinity chromatography.

Biotinylation

Biotinylated through reaction with a N-hydroxysuccinimide ester of biotin.

Concentration1 mg/ml
Supplied in

PBS with 0.02% sodium azide. Sterile-filtered (0.2 µm).

Contents

Monoclonal antibody MT2/4/6, biotinylated. Supplied at 1 mg/ml in PBS with 0.02% sodium azide.

Shipping and Storage

Shipping

Shipped at ambient temperature.

Storage

Store product at 4-8°C or frozen at -20°C or below. Avoid repeated freezing/thawing.

Shelf lifeAt least 18 months from date of receipt.

The systems reactive with NHPs are either based on cross-reactive human kits or specifically developed monkey kits (NHP).

Cross-reactivity verification tests have been performed in ELISpot and/or ELISA by Mabtech and/or by others. Evaluations of assays with less solid evidence of cross-reactivity are shown as symbols within parentheses.

 Rhesus MacaqueCynomolgus MacaquePig-tailed MacaqueBaboon*Sooty MangabeyAfrican Green MonkeyNight Monkey*Common MarmosetSquirrel Monkey
ASSAYOld worldNew world
ApoA1      
ApoB       
ApoE (NHP)       
ApoE       
ApoH       
CCL2 (MCP-1)       
CCL4 (MIP-1β) ( ) ( )( )( )( )
CCL22 (MDC) ( ) ( )( )( )( )
CD25       
GM-CSF( )( )   
Granzyme A        
Granzyme B (NHP)       
Granzyme B     
IFN-α2        
IFN-α pan   ( )   
IFN-γ (NHP)
IFN-γ( )
IgA (NHP)       
IgA       
IgG       
IgM       
INS     
IL-1α        
IL-1β        
IL-2 (NHP)
IL-3       
IL-4( )( )( )( )
IL-5( )( )( ) 
IL-6( )( ) ( ) 
IL-8 (NHP)        
IL-8 (ELISA)        
IL-8 (ELISpot)        
IL-10 (NHP)      
IL-10       
IL-12/-23 (p40)      
IL-12 (p70)        
IL-13 (NHP)( )( )( )  
IL-13        
IL-17A (NHP)( )( )   
IL-17A     
IL-17F        
IL-17A/F        
IL-21      
IL-22        
IL-23     
IL-27        
IL-31        
IP-10       
Perforin( )   
TGF-β1 (latent)       
TNF-α (NHP)( )  ( )( )( )( )
          
SINGLE mAbs         
Cell stimulation         
CD3, mAb CD3-1       
CD3, mAb CD3-2       
CD28, mAb CD28-A       
Flow cytometry          
IFN-γ mAb 1-D1K( )( )( )( )( )( )
IL-2 mAb MT8G10( )( )( )( ) ( )( )( )
IL-4 mAb IL4-3( )       
IL-17A mAb MT504        
Perforin mAb Pf-344     
TNF-α mAb MT15B15       
Ig reagents         
IgG1 mAb MTG1218       
IgG1 mAb MT1939       
IgG2 mAb H6200       
IgG2 mAb MTG211E       
IgG3 mAb MTG34       
IgG4 mAb MTG42       
Western blot         
IFN-γ mAb MT111W( )( )( )( )( )( )( )( )( )
TNF-α mAb MT15B15( )( )( )  ( )( )( )( )

Symbol keys:


Cross-reactivity

Verified by Mabtech and/or by others. Tests performed in ELISpot and/or ELISA.

  • good
  • poor
  • no

Indications of Cross-reactivity

Based on reports from others or by analysis using recombinant proteins.

  • ( ) good
  • ( ) poor
  • ( ) no

Monkey kits

Specifically developed monkey kits are marked with (NHP).

Blank boxes

Represent cross-reactivities not evaluated.

* Comprises several species. Cross-reactivity may have to be verified on a species basis.

Find out which analyte combinations we have evaluated in T cell FluoroSpot and which combinations are affected by capture effects or not. 

What is a capture effect?

When capture antibodies with different specificities are coated together, the capture of one cytokine may affect the secretion of other cytokines. This is usually more pronounced when studying T cell responses with polyclonal stimuli compared to antigen-specific responses. Capture effects are seldom a problem and can often be counteracted by the addition of an anti-CD28 antibody.

Depiction of capture effects observed in FluoroSpot

(1) IL-2 secreted by the activated T cell is captured by coated anti-IL-2 capture antibodies. (2) As a result, IL-2-stimulation of the T cell itself (autocrine stimulation) as well as nearby T cells (paracrine stimulation) is impaired, ultimately leading to (3) decreased IFN-γ secretion.

Co-stimulation with anti-CD28

Anti-CD28 mAb provides a co-stimulatory signal to antigen-specific responses by binding to CD28 on T cells. The addition of an anti-CD28 mAb to the cell culture enhances antigen-specific responses and can counteract capture effects. For example, the presence of IL-2 capture antibodies may result in reduced activation of T cells, as capturing of IL-2 decreases the amount of available IL-2 and thereby dampens the IL-2-mediated stimulation of T cells. The addition of anti-CD28 mAb restores IFN-γ responses (depicted in the below images). Further optimization may be necessary, depending on the cells and stimuli used. Too high concentrations of the anti-CD28 mAb may result in an elevation of non-specific cytokine secretion.

Overcoming capture effects using anti-CD28 anitbodies in FluoroSpot

(1) An anti-CD28 antibody can be added to provide a co-stimulatory signal that can restore (2) e.g. IFN-γ responses.

How to investigate capture effects

Our FluoroSpot Plus kits are evaluated for capture effects, and in studies of T cell responses we recommend co-stimulation with anti-CD28. With FluoroSpot Flex, it is possible to combine and build your own kit. For guidance look at our analyte combination table (above). Capture effects can be investigated by quantifying spot numbers in wells coated with single capture antibodies and compared to wells coated with a mixture of the capture antibodies. The compensatory effect of the anti-CD28 mAb may be assessed by comparing cells cultured with and without the anti-CD28 mAb.

Tutorial, Published December 28, 2023

Incubation strategies for success in ELISpot and FluoroSpot
We've made a handy table summarizing incubation times for different analytes to make your research planning for the ELISpot and FluoroSpot assay a little easier.
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IFN-α pan

Analyte description

Interferon-α and interferon-β are type I interferons. These antiviral cytokines are mainly produced by virus-infected cells after sensing the presence of danger signals such as viral DNA or RNA. IFN-α and IFN-β act through the same receptor and initiate an anti-viral response, affect immune regulation, T cell responses, and also anti-tumor effects have been shown. A key producer of type I interferons is the plasmacytoid dendritic cell. Additionally, IFN-α is secreted by lymphocytes, macrophages, fibroblasts, and dendritic cells. 

In humans, interferon-α has 13 subtypes: IFN-α 1, 2, 4, 5, 6, 7, 8, 10, 13, 14, 16, 17, and 21.

In mice, there are 14 IFN-α subtypes: 1, 2, 4, 5, 6T, 7/10, 8/6, 9, 11, 12, 13, 14, 15 (A), and B. There are also different allelic forms of mouse IFN-α subtypes 1, 7/10, 8/6, and 11 in mouse strains 129/Sv and C57BL/6.

Alternative namesInterferon-α, Interferon-alpha, IFN-α, IFN-alpha, IFN-a, IFNA
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