1. Initial Meeting: Participate in a 30-minute meeting to present the kit and address any questions.
   

2. Kit Shipment: Confirm participation, send a purchase order (PO), and receive the kit. Shipping typically takes less than a week to the USA or Europe.
   

3. Kit Storage and Preparation: Store the kit properly until use. Optionally, request a protocol walk-through meeting to discuss specific applications and procedures.
   

4. Wet Lab and Sequencing: Perform the wet lab procedures and, if you want, send the quality control (QC) on the sample to us to ensure everything is in order. Then proceed with the selected cDNA library preparation and sequencing.
   

5. Bioinformatics Analysis: You can opt for running our standalone proprietary software or ask for a basic analysis with comprehensive report. In the second case, you will receive the bioinformatics report within two weeks. After that, you can participate in a one-hour debriefing session to discuss results.
   

6. Feedback: Complete a feedback form at the end of the program.

The single-cell kit contains two boxes with all necessary reagents for single-cell isolation, barcoding, and cDNA generation. The kit includes:

• Box 1: Cell partitioning chip, magnet, barcoded beads, and wash buffers A and B (stored at 2°C to 8°C).
  

• Box 2: Lysis buffer, RNase inhibitor, DTT, RT master mix, reverse transcriptase mix, TS primer, amplification enzyme, amplification master mix, and A primer mix (stored at -25°C to -15°C).
  

• Bioinformatics data analysis: Access to our standalone proprietary software or a basic bioinformatics report performed in our cloud.

• Dedicated technical support : Direct access to expert guidance on experimental design and analysis.
  

Available in presentations for 2 or 16 reactions.

Note: Sequencing library preparations for PacBio/ONT platforms must be ordered separately.

Specific Target: The magnetic beads specifically target poly-A tailed mRNA by having a poly-T sequence at the 3’-OH end.

* Contact us for customized kit configurations for targeted gene panels.

Number of UMI (unique molecular identifiers) per Bead: Each bead contains 10^7-10^8 unique binding sites, each associated with a unique molecular identifier (UMI). This means each bead can bind to 10^7-10^8 different mRNA molecules, each tagged with a unique UMI.

• Standard Chip Compatibility: The standard chip is compatible with cells or nucleus preparations ranging from 5 to 25 μm in size.
  

• Customized Chip Requirements: Larger cells (up to 70 μm) or irregularly shaped cells require a customized chip.
  

• Sample Preparation Recommendations: Fresh cell samples with high viability in PBS solution are recommended.
  

• Security Features: The chip is disposable, making it suitable for handling high-security samples, such as those used in HIV investigations.
  

• Successfully Tested Cell Types: Examples of compatible cell types include PBMC, ZebraFish embryo cells, HEK-293, NIH-3T3, MDA-MB-453, K-562, A549, and PC3, among others.

• Amount of cell/nuclei:

- Per Chip: Each chip can partition between 3,000 to 10,000 cells by loading 15,000 to 35,000 cells.

- Combined Analysis: Up to 20,000 cells can be analyzed together by pooling two reactions, thanks to different barcodes on the generated cDNA.

The Single-Nucleus RNA Library Kit is under final validation; contact us for beta testing program.

Yes, the chip can be operated manually without any special equipment. You only need a regular P200 pipette to load the chip.

The process from sample to cDNA takes just 5 hours with minimal hands-on time—less than 1 hour of total pipetting—ensuring high reproducibility and low error rates.

The width of the tip is crucial for chip loading, and using the right tip can significantly facilitate loading. Here are some pipette tips that have been validated for successful chip loading:

• Eppendorf cat #0030078551

• Jet Biofil PMT231200

• VWR cat #732-3701

• Vertex #F42L7N0F-VXRG

• Rainin cat#30389242

• Extragene TS250CFS

• Opentrons OT-2 #999-00081
  

For first-time users, it is recommended to use some of these P200 tips.

If you have a different brand of pipettes or need further guidance on tip selection, you can consult with us for advice on the best tip to use for your specific setup.

• Visual Monitoring: The microwell chip is designed as an open system, allowing users to visually monitor the entire process.
  

• Magnification: The process can be observed with a 10X magnification, providing clear visibility of cell and bead loading.
  

• Transparency and Control: The design enables step-by-step observation, offering transparency and control over each stage of the workflow.

Consumables:

• Regular sterile RNAse/DNAse-free certified filter tips

• 1.5 mL nucleic acid low-binding tubes

• PCR tubes
  

Equipment:

• Single channel pipettes

• Cell counter or inverted microscope and hemocytometer

• Mini-centrifuge

• Vortex

• Thermal mixer

• Thermal cycler

• DynaMag™ magnetic rack

• QuBit™ 4 fluorometer or equivalent

• Nucleic acid fragment analyzer

• ONT/PacBio Sequencer
  

Reagents:

• Absolute ethanol

• Nuclease-free water

• RNase AWAY™ or equivalent

• 10 mM Tris-HCl pH 8.0

• 10% Tween-20

• PBS pH 7.4

• AMPure XP or PB

• QuBit™ dsDNA quantitative kit or equivalent

• Library prep and sequencing reagents

General Compatibility: The kit is specifically designed for long-read sequencing platforms and is compatible with both PacBio and ONT technologies. This allows for the use of the same cDNA preparation for both platforms.

* Users can also analyze their cDNAs with Illumina for same-sample long- and short-read comparisons.

Library Preparation Recommendations:

• For PacBio, Kinnex Full Length RNA is recommended for maximized read-length and Kinnex 16S rRNA kit for maximized number of reads.

• For ONT, LSK 114 is recommended.
  

ONT Single-Cell Experiments:

• For single-cell experiments using ONT, it is recommended to use a P2 or superior equipment.

• Data should be basecalled in SUP or HAC mode for optimal results.

With full-length transcript resolution, you can analyze transcript information that is inaccessible to short-read platforms, including:

• Complete isoforms,

• Gene fusions,

• Alternative splicing events,

• Allele-specific expression,

• Complex loci (TCR/BCR/HLA, pseudogenes)

The basic bioinformatics report includes:

• Sequencing and sample statistics

• Pipeline information

• Details on computational resources used

• Overview of read structure (to check for chimeric reads or other artifcats)

• Cell calling metrics

• Seurat plots

• SQANTI3 plots

Please contact us if you are interested in customized data processing.

Data Submission: You will need to provide raw sequencing data for demultiplexing using ArgenTag's proprietary barcode calling software.

Confidentiality: Your data will be kept confidential and not shared or used for unauthorized purposes, as outlined in our terms and conditions.

You can also access to our standalone, user-installable version of our barcode calling software.

ArgenTag demultiplexing pipeline:

• Minimal requirements: x64 Linux, 8 GB RAM or greater, i3 or greater (or equivalent)

• Recommended requirements: 16 GB or greater, i7 or greater (or equivalent), 8+ cores.

ArgenTag single-cell analysis pipeline:

• A standard experiment can be analyzed with the following setup: x64 Linux, 128 GB RAM or greater, i7 or greater (or equivalent), 16+ cores, at least 60 GB of free disk space.

The minimum requirements depend on the sequencing results. Increasing the available RAM and number of cores will significantly reduce processing time.

Request access to our GitHub for a complete installation tutorial.