LeukoComplete™ Gene Detection Kit for human GAPDH

Cosmo Bio

Catalog No.: MIN-11150-HC-002

Primer and Enzyme for Real-Time PCR

LeukoComplete™ Cellular Immunity Detection Kit

This research-use-only kit evaluates changes in gene expression in antigen-stimulated T cells via qPCR. A small amount of whole blood is treated with viral peptides and applied to a dedicated plate for analysis.

Features

This kit allows for the extraction and purification of mRNA from white blood cells and subsequent cDNA synthesis. The resulting cDNA can be used for qPCR to evaluate gene expression changes. It supports the EAGL (Ex vivo Activation of Gene in Leukocytes) method and includes high-throughput plates: Leukocyte Isolation Plate and mRNA Capture Plate.

Specifications

Applications	Cellular immune response assessment, cytokine gene detection
Work time	180 minutes + stimulation time
Sample types	Fresh blood (human/mouse), PBMC (human/mouse), spleen (mouse)
Final product	30 µL cDNA
Detection targets	Control genes: ACTB, GAPDH, B2M Human genes: IFNG, IL-2, TNFSF2, IL-6, IL-10, GM-CSF, CXCL10, IL-4, IL-5 Mouse genes: IFNG, IL-10, IL-17
Instruments	Real-time PCR, Digital PCR
Use cases	Antigen screening, vaccine efficacy, allergy assessment

Kit Components

Plate and RT Kit (Cat. No. 11130)

• PT Buffer (15 mL)
• Leukocyte Isolation Plate
• Deep Well Plate
• Proteinase K (30 µL)
• TCEP (300 µL)
• Lysis Buffer (6 mL)
• mRNA Capture Plate
• Wash Buffer A (30 mL)
• Wash Buffer B (50 mL)
• Aluminum Seal
• M-MLV Reverse Transcriptase (40 µL)
• RNase Inhibitor (10 µL)
• RT Buffer (1.5 mL x 2)

Antigen Coated Plate (Cat. No. 11140)

• SARS-CoV-2 spike peptide antigen plate (24 tests)

* Overlapping 15-mer peptides with 11 aa overlap. Includes blank, two antigen, and positive control wells.

Gene Detection Kit

• Primer Mix (100 µL)
• PCR Enzyme Mix (500 µL)

* Varies by gene of interest.

Product Data

Gene Extraction Performance

Linear gene extraction (R² > 0.9) from various sources including diluted PBMCs and spleen cells. Figure 1

Target Gene Expression Over Time

Expression profiling of IFNG, IL2, IL4, GM-CSF in fresh human blood after stimulation with CEF peptides. Figure 2

Comparison with ELISpot and ICS

Evaluation of 21 vaccinated individuals showed LeukoComplete™ has comparable or better sensitivity to ICS. Figure 3

Comparative Ct Method

Gene fluctuation assessed via ΔΔCt = (Ct_target - Ct_control [stimulated]) - (Ct_target - Ct_control [unstimulated]). Figure 4

Evaluation Conditions for Target Genes

See full table for gene response to various stimulants in human and mouse cells including PMA, PHA, LPS, ConA, CEF.

Abbreviations

• PMA: Phorbol 12-myristate 13-acetate
• IM: Ionomycin
• PHA: Phytohemagglutinin
• LPS: Lipopolysaccharide
• ConA: Concanavalin A
• CEF: CEF peptide pool

Expression Ranking

Rank	Log2FC (n=12)
+++	> 8
++	5–8
+	3–5
+/-	1.5–3
N.D.	< 1.5
-	Not evaluated

References

1. Mitsuhashi M, et al. Clin Chem. 2006;52(4):634-42.
2. Mitsuhashi M. J Immunol Methods. 2010;363(1):95-100.
3. Saito et al. BBRC. 2023. https://doi.org/10.1016/j.bbrc.2023.149398

Note: This is a research reagent. Do not use for diagnostic, therapeutic, or food applications.

Product Specifications

• Reactivity: Human