Frequently Asked Questions

Consult a list of the most frequently asked questions covering technical specifications and experiment best practices.
If you didn’t find what you are looking for, please reach out to our Customer Support Team at customer@immudex.com.

General 

What are Dextramer® reagents?

Dextramer® reagents are MHC multimers optimized to accommodate multiple binding sites and numerous fluorophores to boost avidity in binding antigen-specific CD4+ and CD8+ T cells. Thus, each molecule presents several options to bind cell receptors in a flexible configuration that promotes multi-binding of antigen-specific T cells to isolate populations even when these are rare or have low-affinity receptors.
That same configuration is at the core of other Immudex technologies to enable multiplexing and expand the types of cells that can be targeted to unravel the complexity of the immune response.

Why is it important to look at antigen-specific T cells?

T cells play key effector and regulatory roles in the immune response to external pathogens, allergens, and self-antigens. Therefore, antigen-specificity of the immune cells can help us gain a deeper understanding of the immunology of different diseases and related therapeutics. The antigen-specificity can be measured indirectly through functional assays like chromium release, cytokine production assays (ELISpot or ELISA), or via intracellular cytokine staining. Based on the MHC-peptide affinity for the TCR, Dextramer® staining is a direct measure of antigen-specificity that allows detection of antigen-specific T cells by flow cytometry, in situ, NGS, or single-cell multi-omics. Dive into the world of antigen-specific T-cell detection in our resource section.

Why is it important to have high-avidity reagents when detecting antigen-specific T cells?

As the affinity of isolated, soluble monomeric MHC-peptide complexes for their specific TCR is weak, and the interaction between the MHC-peptide and associated TCR has a half-life of approximately 10 seconds, multimerization is applied to enhance detection sensitivity. The detection sensitivity increases with the amount of MHC monomers complexed onto a multimer. For enhanced detection sensitivity, the Dextramer® technology holds an optimized number of MHC-peptides that enable efficient and sensitive detection of antigen-specific T cells. Read more about the sensitivity of the Dextramer® reagents here.

Please read more about Dextramer® Technology here or contact customer@immudex.com for further inquiries about Dextramer® products. 

What is a Dextramer® reagent or a dCODE Dextramer® reagent staining?

Dextramer® reagent staining is an assay where T cells are labeled with a fluorophore-labeled Dextramer® reagent for analysis by flow cytometry and in-situ staining to detect their antigen-specificity. Find the staining protocol here.

Dextramer® reagents enable you to: 

  • Efficiently detect multiple T-cell specificities in the same sample, whether blood samples, tissue-derived cell samples, or in vitro cell cultures by flow cytometry or next-generation sequencing
  • Combined staining with other cell surface markers or intracellular cytokine or chemokine staining for a more comprehensive characterization of antigen-specific T cells

  • Combined with single-cell analysis platforms, such as 10x Chromium or BD Rhapsody™ systems, dCODE Dextramer® reagent allows for characterization of T-cell antigen specificity at single-cell resolution, providing simultaneous information on gene expression, VDJ sequences, and surface marker expression. Read more about dCODE Dextramer® reagents here

Can I store Dextramer® reagents or dCODE Dextramer® reagents in the freezer?

No, freeze-thaw cycles of Dextramer® products will decrease or completely eradicate staining efficacy and is, therefore, not recommended. 

Can I fix my cells before staining with Dextramer® reagents?

For optimal MHC Dextramer® reagent staining of antigen-specific T cells, the staining should be performed on live cells. However, if you wish to analyze your cells later or you are staining for intracellular markers, Dextramer® reagent staining should be performed before fixation. You may also want to consider additional washing steps before proceeding to intracellular staining. We recommend using the following reagents: IntraStain (Dako, cat no. K2311), BD Cytofix/Cytoperm™ (BD, cat no. 554714), or Cyto-Fast™ Fix/Perm (BioLegend, cat no. 426803). 

What is the stability/shelf life of MHC Dextramer® reagents?

The shelf life for each Dextramer® product will vary. In general, the higher affinity of the peptide to the MHC, the more stable the MHC Dextramer® reagent will be, resulting in longer expected shelf life. Therefore, each MHC Dextramer® reagent will have its own unique shelf life, and a general shelf life for all MHC Dextramer® reagents cannot be provided. One way to assess the status of your Dextramer® reagent is to store a reference sample that can be used to check the status of your MHC Dextramer® reagent over time.

Why is MHC-matching important? / Why does the MHC type of my cells have to be the same as the MHC on the Dextramer® reagent?

MHC-matching is essential to ensure a biologically relevant read-out. Each donor expresses two MHC I and MHC II alleles, i.e., 2 sets of 3 MHC I genes and two sets of 3 MHC II genes. The immune system is highly specific to each individual and the antigen-specific T cells of that donor will, therefore, only detect a given peptide presented by MHC molecules expressed in said donor.

How do I identify the optimal peptide?

MHC class I molecules typically bind 9-mer peptides, but other peptide lengths may be relevant. Especially for MHC I, it is important to identify the optimal peptide you wish to bind to the MHC molecule of interest. There are a few peptide predicting tools online:

http://www.cbs.dtu.dk/services/NetMHC/ (recommended)

http://www.syfpeithi.com/ 

If you need help in choosing the correct peptide sequence, our experts are always ready to help. Contact us at customer@immudex.com  

What quality of peptide do I need for loading?

At Immudex®, we recommend a level of peptide purity of 95% when loading easYmer® and U-Load® MHC II monomers as loading of peptides with lower purity can be difficult and compromise the quality of the detection of antigen-specific T cells.

Controls

Why are negative controls required? What would be a good negative control for T cells or B cells?

Using one or more negative controls will increase the chance of unequivocal results and help set the proper cut-off between negative and positive populations. They help discriminate real events from artifacts, allowing the selection of a background threshold that will help you define your true positive cells.

You may choose an allele-matched or an allele-mismatched negative control:

  • Allele-matched control: same allele as your reagent but a different and irrelevant peptide
  • Allele-mismatched control: MHC of a different allele than expressed by the donor you are to analyze and a different peptide

We have four negative controls carrying non-sense peptide sequences for Human MHC I. Those can be manufactured both as MHC Dextramer® reagents or dCODE Dextramer® reagents.

Available Negative Controls at Immudex ® 

Human MHC I 

  • HLA-B*0801/AAKGRGAAL (cat# WI03233/NI03233) 
  • HLA-A*0201/ALIAPVHAV (cat# WB02666) 
  • HLA-A*0101/STEGGGLAY  
  • HLA-A*0301/GLFGAGAFK 

Human MHC II 

  • DRB1*0401/ PVSKMRMATPLLMQA (CLIP peptide) (cat# FB10002)
  • DRB1*0402/ PVSKMRMATPLLMQA (CLIP peptide) 
  • DRB1*1101/ PVSKMRMATPLLMQA (CLIP peptide) (cat# FD10002)
  • DRB1*0701/ PVSKMRMATPLLMQA (CLIP peptide) 
  • DRB1*0301/ PVSKMRMATPLLMQA (CLIP peptide) 
  • DRB1*1301/ PVSKMRMATPLLMQA (CLIP peptide) 

Mouse MHC I 

  • H2-Kb / SIYRYYGL (SIY) (cat# JD02164)*

*H2-Kb / SIYRYYGL (SIY) can be used as a positive and negative control depending on the mouse model. Contact customer@immudex.com for more information. 

Mouse MHC II 

No pre-designed controls (see recommendations below) 

Alternatively, you can design a Dextramer® reagent carrying a peptide epitope unrelated to your experimental system and for which you would not expect to find any antigen-specific T cells. The peptide-MHC binding affinity must be high enough to manufacture a stable Dextramer® reagent; therefore, we screen the peptide-MHC combination in a prediction software before manufacturing. (NB: this is what we would recommend for MHC Class II Dextramer® reagent as we have no pre-designed negative controls besides the CLIP reagents). Other relevant controls could be a negative donor sample for the specific antigen in question or an allele-mismatched donor.

Learn more about the importance of Positive and Negative controls here

When is a positive control necessary? What would be a good positive control for T cell and B cell assays?

A positive control can help you ensure that your experimental set-up works properly and can help you optimize your staining and your general workflow. "Positive controls" can be divided into two groups:

  1. A cell control: for example, a sample with a known frequency of antigen-specific T cells detected by your defined MHC Dextramer® reagent
  2. An assay control: It could be a Dextramer® reagent that recognizes very common epitopes in the donor of interest. We have performed a study showing that staining HPBMC with these 3 MHC Dextramer® reagent specificities gives coverage of 95% of A*0201 restricted donor samples i.e., 15/16 of the donor samples were positive for one or more of these three specificities.

Suggested Positive Controls at Immudex®  

Human MHC I 

  • CMV Dextramer® reagent: HLA- A*0201/NLVPMVATV  
  • EBV Dextramer® reagent: HLA-A*0201/GLCTLVAML  
  • Flu Dextramer® reagent: HLA- A*0201/GILGFVFTL  

Learn more about the importance of Positive and Negative controls here

 MHC Dextramer® reagents

Can I stain the same sample with multiple MHC Dextramer® reagents by flow cytometry?

Yes, there are several possible options:

  • For staining of MHC Dextramer® reagents of the same specificity but with different fluorophores, see this protocol
  • For staining of MHC Dextramer® reagents of different specificities, see this protocol

Klickmer® reagents

Why does the ligand have to be mono-biotinylated?

If multiple biotin molecules are present, several Klickmer® molecules can aggregate and may, in turn, precipitate or result in an artifact in your staining. 

What is a good negative control for Klickmer® reagents?

As a negative control, you could load onto the Klickmer® molecule a non-relevant ligand like your ligand of interest (same size, same chemical properties, for instance), but that does not bind the cells you are attempting to identify. Please note that an empty Klickmer® molecule is not an appropriate negative control.

What is the recommended loading range for Klickmer® reagents?

This will vary depending on the ligand of interest and the affinity of the ligand for its target. We, therefore, recommend performing a titration to determine the optimal ligand/Klickmer® reagent ratio for your set-up. Please note that the different colored Klickmer® products have different loading capacities.

Does the assembled Klickmer® molecule need to be diluted for flow cytometry?

Yes, the stock concentration of Klickmer® molecule is 160 nM, and once the ligand is loaded, we recommend diluting the assembled reagent down to 32 nM for flow cytometry staining. 10 µl of the pre-diluted reagent can be used to stain 50-100 µl lymphoid cells (up to 1 x 106 lymphoid cells, or up to 200-500.000 clonal cells).

How long can you store Klickmer® reagents or a loaded Klickmer® reagents?

Klickmer® molecule is stable for two years, but because we cannot foresee all applications of Klickmer® reagents and what ligands will be loaded, we cannot make any claims on the stability for a loaded Klickmer® reagent.

dCODE Dextramer®

Setting up a panel with dCODE Dextramer® reagents

Is there a limit to how big the panel can be?

There is no staining limit in theory, and 150 to up to 1200 specificities were successfully multiplexed in previous projects using dCODE Dextramer® reagents (see Viborg et al. 2019, Pedersen et al. 2019, Bentzen et al. 2016). Note that those used dCODE Dextramer® (HiT) reagents for bulk analysis. In practice, this must be optimized, and one should be careful not to dilute the cell sample too much. We recommend staining with 2 µl for each specificity, and for large panels, we also suggest increasing the incubation time.

Can MHC Class I and II Dextramer® reagents be included in the same panel?

Yes, the incubation time with the dCODE Dextramer® reagent mixture should follow the MHC II Dextramer® reagent staining recommendation, which is 20 min. Also, a 2-way sorting CD4/CD8 must be set up to separate your cells of interest.

Can dCODE Klickmer® reagents and dCODE Dextramer® reagents be run in the same panel?

Yes, in that case, the incubation time with the dCODE Dextramer® mixture should follow the recommendation for MHC I Dextramer® reagent staining, which is 10 min. 

Choosing the right product (dCODE Dextramer® reagent) 

What is dCODE Dextramer® (HiT) reagent, and when should it be used?

dCODE Dextramer® (HiT) reagents are DNA-tagged Dextramer® reagents designed for Epitope Discovery and Neo-antigen Screening. The DNA barcode allows detection of several specificities in the same sample by Next-Generation Sequencing (compatible with several technologies, including Illumina and Ion Torrent sequencing). This approach is ideal for screening a large number of potential epitopes and quickly reducing it to a shorter list of "hits".

What is dCODE Klickmer® reagent and when should it be used?

dCODE Klickmer® reagent is a DNA-tagged Klickmer® reagent designed to accommodate your needs not covered by our standard product lines. It's a flexible solution for NGS (HiT), multiplexing, and single-cell analysis (RiO, 10x) for customers looking at alleles we do not have in stock (with their monomers) or at cell types other than T cells. Any biotinylated molecule can be attached to a dCODE Klickmer® reagent. This multipurpose tool serves a range of applications and can help address diverse needs in research and development:

  • Highly sensitive detection of antigen-specific B cells and other immune cells
  • Identification of ligands for low-affinity interactions
  • Isolation of specific cell populations

dCODE Dextramer® (10x) reagents 

Can two cells be found in the same GEM?

Two cells may be encapsulated in one gel bead, and this possibility follows a Poisson distribution. Therefore, a low number of cells should be used (10.000 cells) to limit the risk of getting two cells in one droplet while the gel beads are added in excess. Due to the Poisson distribution, there is less than 10% chance of having two cells on one gel bead, and therefore in one GEM. The presence of dCODE Dextramer® reagents does not increase the formation of cell doublets. We have observed that the number of doublets is constant with different amounts of dCODE Dextramer® reagents and is comparable to the cells not stained with dCODE Dextramer® reagents.

How many dCODE Dextramer® reagents bind to one cell?

We have not determined this accurately, but our best estimate is up to 2000 molecules per cell. Even two clonal cells can bind different numbers of dCODE Dextramer® reagents depending on their activation status.

Can more than one dCODE Dextramer® molecule specificity bind to one cell/ Can two dCODE Dextramer® molecules with different specificities bind to the same cell?

Yes, it is a possibility, but its signal would be very low, so it will likely not be detected. This will be the case if a TCR clonotype is able to bind dCODE Dextramer® reagents with different specificities or if a cell expresses more than one TCR clonotype. If true cross-binding is not suspected, one can set up a threshold of one dCODE Dextramer® reagent per cell in the analysis to filter out cells recognized by multiple specificities and reduce background.

After deconvolution of data, do you still retain information about the number of dCODE Dextramer® reagent bound to a given cell?

Yes, you can identify how many dCODE Dextramer® molecules are bound to a given cell by looking at the UMI. Up to 2000 dCODE Dextramer® molecules can bind. Still, the exact number is highly dependent on the specificity/affinity of the T cell towards the relevant pMHC as well as the T cell activation status and the experimental settings. 

If a dCODE Dextramer® reagent binds to only one cell, can I trust this to be an actual hit and not background? 

Yes, the resolution of the 10x workflow is designed to detect even one cell, and we do have experience with single-cell clones. This frequently occurs in naïve T cells and for certain antigens. Cancer antigens, for instance, are more likely to have single-cell clones than viral antigens. You can see the difference in this case study, for example (dCODE Dextramer® (10x compatible) Case Study), where out of 187 cells specific to the cancer antigen MART-1, 180 different clonotypes were identified. Of course, multiple cells should ideally define a population, but detecting one cell is not uncommon and should not be systematically regarded as background.

How do you define "specific binders "?

Some general rules to define specific binders are:

  • The dCODE Dextramer® reagent-positive antigen-specific T-cell populations should have a signal over the threshold defined by the general staining of the negative control reagents.
  • A positive cell should be of the right cell type, i.e., CD8 or CD4-positive.
  • Only MHC matching the alleles of the donor should be considered positive. 
  • Antigen-specific T cells binding more than one dCODE Dextramer® reagent should generally be excluded (except in cases where genuine cross-reactivity is suspected)

However, please consider that MHC-peptide complexes are complex molecules that can have "specific" interactions with non-antigen-specific T cells. Therefore, it is possible to find cells interacting with the dCODE Dextramer® reagent with a signal above the background but are not antigen-specific cells. Identifying true antigen-specific T cells relies not only on the dCODE Dextramer® reagent but also on other parameters, such as the right surface markers and expression of T-cell-related genes.

Is it possible to identify the antigen that matches a given TCR CDR3 sequence?

Assessing the specificity of a TCR based solely on CDR3 sequences is very difficult. However, our dCODE Dextramer® technology allows you to test a given T cell with a TCR of interest against any array of potential/predicted pMHC targets. The dCODE Dextramer® technology (10x and RiO) is designed specifically with this in mind.

How many cells can be loaded on a 10x Chromium chip? What is the minimum number that can be loaded?

It is recommended to load between 500-10.000 cells on the Chromium chip per well. The minimum number that should be loaded is 500 cells. Loading more cells is possible, but it increases the chances of cell doublets, i.e., two cells landing in the same GEM.

Is dCODE Dextramer® (10x) reagents compatible with TotalSeq™-C antibodies from BioLegend? How can they be used in the same workflow?

Yes, TotalSeq™-C antibodies can be used like any other antibody following our standard staining protocol after the dCODE Dextramer® reagent staining step.

Can dCODE Dextramer® (10x) reagent and TotalSeq™-C antibodies have overlapping barcodes?

No, dCODE Dextramer® (10x) reagents and TotalSeq™-C antibodies have been assigned separate barcodes, so overlap is not an issue.

Where do I find information on the 10X feature barcodes?

On page 85 in the Appendix of this manual: 10x Chromium Next GEM User Guide

Does including negative or positive controls dilute the signal of interest?

No, it is generally not an issue. However, the positive control should be chosen so that it is not too abundant (frequency below 50%), and a pilot staining should be performed to ensure this is not the case. Alternatively, one can spike in a known number of cells positive for another epitope to have an internal reference for each sample.

dCODE Dextramer® (RiO) ​reagents 

How many cells can be loaded onto a cartridge?

The cartridge contains 200.000 wells, but we recommend loading between 500-20.000 cells. Loading more cells is possible, but it increases the number of multiplets per well (multiple cells landing in the same well) following a Poisson distribution.

Is dCODE Dextramer® (RiO) reagent compatible with AbSeq antibodies?

Yes, BD® AbSeq™ antibodies can be used just like any other antibody following our standard staining protocol after the dCODE Dextramer® reagent staining step.

Can two beads end up in the same well?

No, the beads are loaded in excess (600.000 beads), but the well-size only allows for one bead per well.

easYmer® and U-Load® MHC II monomers

What is an easYmer® and U-Load® MHC II monomer?

easYmer® and U-Load® MHC II monomers are "empty" MHC I or II molecules. The proprietary formulation renders the highly active and peptide-receptive MHC I or II molecules ready for loading and for any non-expert user to produce MHC monomers easily. These monomers can be used to investigate the specific peptide binding affinity or be attached to a U-Load Dextramer® reagent by following a simple protocol.

Read more about easYmer® and U-Load® MHC II monomers here, or download easYmer® or U-Load® MHC II monomers assembly protocol here and here, respectively. Please contact customer@immudex.com for further inquiries about easYmer® or U-Load® MHC II monomers.

Do I need special equipment to assemble my easYmer® or U-Load® MHC II monomers for loading on U-Load Dextramer® reagent?

No, there are no special equipment requirements to set up an easYmer® or U-Load® MHC II monomer loading. All buffers and other reagents needed for folding and loading peptide onto U-Load Dextramer® reagents are provided upon purchase. Simply attach your own antigen to an exceptional selection of MHC allotypes and load the complex onto U-Load Dextramer® reagents or U-Load dCODE Dextramer® reagents.

Read more about easYmer® and U-Load® MHC II monomers here, or download easYmer® or U-Load® MHC II monomers assembly protocol here and here, respectively. Please contact customer@immudex.com for further inquiries about easYmer® or U-Load® MHC II monomers.

U-Load® Technology

What is U-Load Dextramer® reagents?

A U-Load Dextramer® reagent is a Dextramer® reagent empty of monomer and peptide ready for you to load with the combination that you prefer for your research. U-Load Dextramer® reagents can be loaded with an optimal amount of easYmer® or U-Load® MHC II monomer and the specific peptides of your choice. Thereby, U-Load Dextramer® reagents can be designed to detect either CD8+ and CD4+ T cells that express the cognate T-cell receptor (TCR) interacting with the specific MHC-peptide I or II complex. For cells with few receptors on the surface, bright reagents are essential in resolving these dim cells from others in a sample. Therefore, for effective identification of antigen-specific T-cell populations, U-Load Dextramer® reagents can be provided labeled with FITC, PE, or APC.

Please read more about U-Load Dextramer® reagents here or contact customer@immudex.com​ for further inquiries about U-Load Dextramer® reagents.

What is U-Load dCODE Dextramer® reagents?

U-Load dCODE Dextramer® reagent is a dCODE Dextramer® reagent empty of monomer and peptide ready for you to load with the combination that you prefer for your research. U-Load Dextramer® reagents can be loaded with an optimal amount of easYmer® or U-Load® MHC II monomer, provided by Immudex®, and specific peptides for multiplexed screening or single-cell multi-omic analysis of antigen-specific T cells. U-Load dCODE Dextramer® reagent can be used to detect either CD8+ or CD4+ T cells that express the cognate T-cell receptor (TCR), interacting with the specific MHC I or II-peptide complex placed on the dextran backbone. U-Load dCODE Dextramer® reagent has DNA barcodes unique to each T-cell specificity.

As samples here can be massively multiplexed, U-Load dCODE Dextramer® reagent can be provided labeled with the PE fluorophore. The PE fluorophore enables efficient enrichment of relevant antigen-specific T-cell populations by fluorescence-activated cell sorting (FACS). If staining with antibodies for flow cytometry followed by AbSeq or TotalSeq antibodies for sequencing, it is important to remember to choose different clonotypes of the antibodies to ensure high quality binding and eliminate background.

The U-Load dCODE Dextramer® reagent is provided in three different formats:

  • U-Load dCODE Dextramer® (HiT) reagent: Multiplexed screening of antigen-specific T cells by PCR and NGS.
  • U-Load dCODE Dextramer® reagent compatible with the 10x Chromium System (10x) or BD Rhapsody system (RiO): Single-cell multi-omics analysis using 10x Chromium or BD Rhapsody system

Please read more about U-Load dCODE Dextramer® reagents here or contact customer@immudex.com for further inquiries about U-Load dCODE Dextramer® reagents.

Why use U-Load Dextramer® reagent or U-Load dCODE Dextramer® reagent?

U-Load Dextramer® reagent and U-Load dCODE Dextramer® reagent is a novel reagent concept that allows you to combine custom MHC-peptide allele complexes with the benefits of Dextramer® technology. easYmer® MHC I or U-Load® MHC II monomers can be attached to the empty U-Load Dextramer® reagent or U-Load dCODE Dextramer® reagent backbone provided by Immudex® from your choice of a broad range of MHC I and MHC II alleles and your antigen peptides. Thereby, U-Load Dextramer® reagent or U-Load dCODE Dextramer® reagent can be used to easily assemble quality reagents with a simple protocol that takes 1 hour and can be done at your workbench for screening of multiple specificities. U-Load Dextramer® reagent and U-Load dCODE Dextramer® reagent give you the flexibility to choose which specificities to screen and how many tests you need to do so. With its easy and efficient workflow and reduced cost, U-Load Dextramer® reagent and U-Load dCODE Dextramer® reagent minimizes time and money spent to screen large numbers of specificities and samples.

Please read more about U-Load® technology here, find the U-Load® technology assembly protocols here, or contact customer@immudex.com for further inquiries about U-Load Dextramer® reagents or U-Load dCODE Dextramer® reagents.

What is included when purchasing U-Load Dextramer® reagent Kit or U-Load dCODE Dextramer® reagent

The U-Load Dextramer® reagent Kit with easYmer® reagent includes:

U-Load Dextramer® reagent:

  • Fluorescently labeled U-Load Dextramer® reagent (PE, APC, FITC)
  • Dilution Buffer: PBS, 0.5% BSA, 7.5 mM NaN3, pH 7

easYmer® reagent:

  • 3 µM Peptide loadable MHC monomers consisting of peptide receptive, biotinylated MHC monomers in Tris/Maleate pH 7, 30% Glycerol. Each easYmer® reagent is uniquely identified by the allele, e.g., easYmer® HLA-A*01:01.
  • easYmer® reagent Loading Buffer: 0.3 M Tris/Maleate pH 7
  • easYmer® reagent Control Peptide: Positive control for evaluation of peptide-MHC folding. Each easYmer® reagent Control Peptide is uniquely identified by allele and peptide sequence, e.g., easYmer® reagent HLA-A*0101 Control Peptide KSEYMTSWFY. Prior to use, dissolve the peptide in 20 µL DMSO to a peptide concentration of 1 mM

The U-Load Dextramer® reagent kit with U-Load® MHC II reagent, includes:

U-Load Dextramer® reagent:

  • Fluorescently labeled U-Load Dextramer® reagent(PE, APC, FITC)
  • Dilution Buffer: PBS, 0.5% BSA, 7.5 mM NaN3, pH 7

U-Load® MHC II reagent:

  • 16.6 µM Peptide loadable MHC monomers consisting of peptide receptive, biotinylated MHC monomers in PBS, 15 mM NaN3. Each U-Load® MHC II reagent is uniquely identified by the allele, e.g., U-Load MHC II HLA-DRB1*01:01 reagent.
  • U-Load® reagent MHC II Loading Buffer: Phosphate Buffer, pH 5.9
  • U-Load® reagent MHC II Peptide Loading Component: lyophilized

The U-Load dCODE Dextramer® reagent with easYmer® reagent, includes:

U-Load Dextramer® reagent:

  • Fluorescently labeled U-Load Dextramer® reagent (PE)
  • Dilution Buffer: 1xPBS, 0.5% BSA, 7.5 mM NaN3, pH 7

easYmer® reagent:

  • 3 µM Peptide loadable MHC monomers consisting of peptide receptive, biotinylated MHC monomers in Tris/Maleate pH 7, 30% Glycerol. Each easYmer® reagent is uniquely identified by the allele, e.g., easYmer® reagent HLA-A*01:01.
  • easYmer® reagent Loading Buffer: 0.3 M Tris/Maleate pH 7
  • easYmer® reagent Control Peptide: Positive control for evaluation of peptide-MHC folding. Each easYmer® reagent Control Peptide is uniquely identified by allele and peptide sequence, e.g., easYmer® reagent HLA-A*0101 Control Peptide KSEYMTSWFY. Prior to use, dissolve the peptide in 20 µL DMSO to a peptide concentration of 1 mM

The U-Load dCODE Dextramer® reagent with U-Load® MHC II reagent:

U-Load dCODE Dextramer® reagent:

  • Fluorescently labeled U-Load Dextramer® reagent(PE, APC, FITC)
  • Dilution Buffer: 1xPBS, 0.5% BSA, 7.5 mM NaN3, pH 7

U-Load® MHC II reagent:

  • 16.6 µM Peptide loadable MHC monomers consisting of peptide receptive, biotinylated MHC monomers in PBS, 15 mM NaN3. Each U-Load MHC II reagent is uniquely identified by the allele, e.g., U-Load MHC II HLA-DRB1*01:01.
  • U-Load® reagent MHC II Loading Buffer: Phosphate Buffer, pH 5.9
  • U-Load® reagent MHC II Peptide Loading Component: lyophilized

Please read more about U-Load dCODE Dextramer® reagents here or contact customer@immudex.com for further inquiries about U-Load dCODE Dextramer® reagents.

What is the difference between a U-Load Dextramer® reagent and ready-to-use MHC Dextramer® reagents?

MHC Dextramer® reagents are purchased as ready-to-use and loaded with the MHC-peptide complex once it leaves our labs. The MHC Dextramer® reagents provide outstanding detection of antigen-specific T cells. All Immudex® products are produced to the highest standards, resulting in reproducible monitoring over time. Clinical-Grade and IVD Dextramer® reagents are available and currently in use within the USA and Europe. Read more about the quality of production of our reagents here.

U-Load Dextramer® reagents are Do-It-Yourself (DIY) Dextramer® reagents applicable for flexible multimer stainings. U-Load Dextramer® reagent is provided empty of MHC monomer and peptides ready for you to load in the comfort of your lab with the combination that you prefer for your research.

Please read more about U-Load Dextramer® reagents here or contact our customer services on customer@immudex.com to learn more about your options if you need flexible detection of low frequency or low-affinity antigen-specific T cells.

What is the difference between a U-Load dCODE Dextramer® reagents and ready-to-use dCODE Dextramer® reagents?

dCODE Dextramer® reagents are purchased as ready-to-use and loaded with the MHC-peptide complex once it leaves our labs. All Immudex® products are produced to the highest quality standards, resulting in reproducible monitoring over time.

U-Load dCODE Dextramer® reagent is a DNA barcoded version of the U-Load Dextramer® reagents. U-Load dCODE Dextramer® reagent is provided empty of MHC monomer and peptides ready for you to load in the comfort of your lab with the combination that you prefer for your research. Thereby, U-Load dCODE Dextramer® reagents are Do-It-Yourself (DIY) dCODE Dextramer® reagents with the same DNA barcoding system as the classical dCODE Dextramer® reagent. However, with U-Load dCODE Dextramer® reagent you decide what volume and which specificity to produce your reagent in and to. U-Load dCODE Dextramer® reagent applicable is for flexible large-scale screening (HiT) or single-cell analysis of antigen-specific T cells (10x and RiO). Read more about U-Load dCODE Dextramer® technology here.

Contact our customer services on customer@immudex.com to learn more about your options if you need detection of low frequency or low-affinity antigen-specific T cells using U-Load dCODE Dextramer® reagent.

What is the difference between a Klickmer® reagent and a U-Load Dextramer® reagent?

Both Klickmer® reagents and U-Load Dextramer® reagents are based on the Dextramer® technology and holds acceptor sites for loading of biotinylated molecules or MHC monomers, respectively. Furthermore, a key difference is that with U-Load Dextramer® reagent and U-Load dCODE Dextramer® reagent you gain access to the most extensive allele portfolio as of our collaboration with immunAware on the loadable market. Additionally, whereas U-Load® technology is a loadable technology optimized for extensive detection of antigen-specific T cells across world populations, Klickmer® reagent is optimized for detection beyond T cells (e.g., B cells, NK cells, tumor cells, etc.).

Read more about Klickmer® technology here or U-Load® technology here.

What are the benefits of using U-Load Dextramer® reagents and U-Load dCODE Dextramer® reagents?

  • Create fit-for-purpose immune monitoring reagents in your lab
  • All the benefits of Dextramer® technology
  • Access the broadest human allele coverage on the market
  • Eliminate high multimer costs when performing multiplexed screening of antigen-specific T cells
  • Simplify your work: no UV radiation and easy peptide loading
  • Keep your epitope sequence proprietary as there is no need to disclose the sequence

Please read more about U-Load® technology here, find the U-Load Dextramer® technology assembly protocols here, or contact customer@immudex.com for further inquiries about U-Load Dextramer® reagents or U-Load dCODE Dextramer® reagents.

How long can I store the easYmer® monomers and U-Load Dextramer® reagents?

The easYmer® monomers are supplied in a stabilized peptide receptive conformation which allows storing in the freezer (-20°C) until expiry date as indicated on the product label. The shelf life for each MHC-peptide loaded U-Load Dextramer® product will vary. The higher affinity of the peptide for the cognate MHC is the more stable the U-Load Dextramer® reagent - and longer shelf life can be expected. Therefore, each loaded U-Load Dextramer® reagent has its own unique shelf life, and we cannot provide general shelf life for loaded U-Load Dextramer® reagents. Accordingly, there is no expiration date on our empty research-use-only U-Load Dextramer® products.

Please read more about U-Load Dextramer® technology here, find the U-Load® technology assembly protocols here, or contact customer@immudex.com for further inquiries about U-Load Dextramer® reagents or U-Load dCODE Dextramer® reagents.

Are the MHC monomers for U-Load Dextramer® reagents quality controlled, and can I control loading myself?

All easYmer® MHC I monomers are functionally tested by immunAware while the U-Load® MHC II reagents are QC tested by Immudex®. The U-Load Dextramer® kit is a complete kit to enable safe and easy assembly and an allotype-relevant positive control peptide is provided with the easYmer®, allowing you to validate your peptide-HLA complexes' quality in a simple flow cytometry-based assay in your lab.