dsDNA IgG ELISA Kit

Name: dsDNA IgG ELISA Kit
SKU: DEIA277
Rating: 5 (1 reviews)

Regulatory status: For research use only, not for use in diagnostic procedures.

COA

SDS

Datasheet

Summary

Sample

Serum and plasma

Species Reactivity

Human

Detection Method

iELISA

Intended Use

The present enzyme-linked immuno sorbent assay (ELISA) is intended for the quantitative or qualitative determination of IgG antibodies directed against dsDNA in human serum or plasma. The immobilised antigen is a highly purified preparation of plasmid dsDNA (90% in supercoiled state ) free from chromosomal DNA and protein (histones). The test is fast (incubation time 30/ 30/30 minutes) and flexible (divisible solid phase, ready-lo-use reagents). Six calibrators allow quantitative measurements; a negative and a positive control check the assay performance.

Contents of Kit

1. MTP 1 Microtiter Plate, coated with dsDNA and hermetically packed in a foil laminate pouch together with a desiccant bag. The plate consists of 12 strips, each of which can be broken into 8 individual wells.
2. ENZCONJ IgG Enzyme Cconjugate IgG, 14 mL, ready-to-use, red coloured. Buffered solution containing stabilising protein, methylisothiazolone and bromonitrodioxane.
3. CAL A-F 6 Calibrators, 2.0 mL each, 0 - 6.5 - 16 - 40 - 100 and 250 IU dsDNA IgG / mL, ready-to-use, gradually blue coloured. Contain TBS, BSA, Tween and Na-azide.
4. CONTROL -and CONTROL + Negative and Positive Control, 2.0 mL each, ready-to-use, green and red coloured, respectively. Contain TBS, BSA, Tween and Na-azide.
5. SAMPLEDIL Sample Diluent, 100 mL, ready-to-use, orange coloured. Contains Tris-buffered saline (TBS), bovine serum albumin (BSA), Tween and Na-azide.
6. TMB SUBS TMB Substrate Solution, 14 mL, ready-to-use, colourless. Contains a buffered solution of TMB and H₂O₂. Contained in a vial impermeable to light.
7. WASHBUF CONC Wash Buffer, 100 mL, 10×-concentrate, blue coloured. Contains TBS, Tween and bromonitrodioxane.
8. STOP TMB Stop solution (0.2 M H₂SO₄), 14 mL, colourless, ready-to-use. Caution: sulfuric acid is corrosive.
9. Directions for use
10. Lot-specific certificate of analysis

Storage

Store kit at 2-8°C. It is stable up to the expiry date stated on the label of the box. Do not use kit beyond its expiry date.

Performance Characteristics

1. Standardisation
The test is standardised with a purified mAb preparation containing IgG antibodies specifically directed at dsDNA. This preparation has been calibrated against the first international standard for dsDNA-Ab, coded Wo/80. The degree of sample reactivity is measured in international units (IU dsDNA IgG / mL).
2.Homogeneity of the solid phase
Measurement of the solid phase homogeneity is a regular QC part of each production lot. This is determined by 288-fold measurement of a positive but non-saturating sample on 3 selected plates. Acceptance criterion: mOD-coefficient of variation (cv) over the plates < 8%.
The figure below shows a representative excerpt (solid phase lot no. 2210R) of such an analysis.

Precision

For the assessment of the test precision, the variability of results under the following conditions was determined: a. within 1 assay and between 3 assays, b. between 3 operators and c. between 2 kit lots.
a. Intra- and inter-assay variability (n = 24 and 72, respectively)

b. Operator to operator variability (n = 12)

c. Variability between 2 kit lots (n = 6)

Detection Limit

The detection limit is defined as that concentration of analyte that corresponds to the mean absorbance of Sample Diluent plus 3-fold standard deviation (s). It was determined as < 1 IU dsDNA IgG per mL sample (n = 24).
Recommended measuring range: 5 – 250 IU dsDNA IgG per mL sample

General Description

Systemic lupus erythematosus (SLE) is an autoimmune-mediated, chronic inflammatory illness with variable clinical presentation, ranging from localised skin lesions to a destructive systemic disorder without cutaneous changes (1). Antibodies to double-stranded (ds)DNA are a well-known, specific marker for SLE with a prevalence from 50-90%, depending on disease severity (2, 3, 4). Their titre often correlates with disease activity (5). Circulating Anti-DNA/-DNA immune complexes are considered to play a role in the pathogenesis of SLE (6). dsDNA-antibodies constitute a SLE criterion according to the American Rheumatism Association (ARA).

Citations

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Background

Systemic lupus erythematosus (SLE) is a systemic autoimmune disease characterized by abnormal activity of the immune system with a wide range of clinical manifestations including renal, cutaneous, neuropsychiatric and cardiovascular symptoms. Biomarkers play a crucial role in the diagnosis of SLE, classification of SLE complications, assessment of SLE disease activity, and reflection of the efficacy of SLE interventional therapy. The diagnosis and classification of SLE is based on the patient's clinical symptoms, signs and laboratory biomarkers reflecting immune reactivity and inflammation in various organs. However, because SLE can cause multi-organ damage, complex pathogenesis, and heterogeneity of clinical manifestations, a particular biomarker may reflect only a specific aspect of SLE, but not the overall state of the disease.

Biomarkers commonly used to assess SLE Figure 1. Common biomarkers for SLE and their measurement sites in patients with SLE
(Source: Yu H, et al. 2021)

Defective clearance of apoptotic and necrotic cells as well as defective degradation of NETosis (neutrophil extracellular traps) may play a crucial role in SLE, where autoantigens contain nuclear material released from the dead cells described above, which are exposed to antigen-presenting cells (APCs), processed by the APCs, and then presented to the autoreactive T cells or B cells. In this process, dysregulation of multiple immune pathways, secretion of pro-inflammatory cytokines, autoantibody production and activation of the complement system ultimately lead to loss of autoimmune tolerance. Anti-dsDNA antibody is one of the most widely used SLE biomarkers and an organ-specific SLE biomarker that assesses, monitors, or predicts organ-specific damage in SLE patients, especially with strong correlation to lupus nephritis, which contributes to organ-specific precision medicine in SLE patients. Anti-DNA antibody binds directly to DNA antigen or cross-reactive antigen in renal cells and indirectly forms an immune complex containing anti-dsDNA antibody in the renal parenchyma, which triggers a complement cascade reaction accompanied by infiltration of immune cells and release of cytokines, thereby inducing renal inflammatory and fibrotic processes.

Anti-dsDNA antibodies are associated with SLE disease activity, so their levels fluctuate over time. Anti-dsDNA antibodies may disappear during treatment but reappear during flares, especially in active nephritis. Anti-dsDNA antibodies are highly specific for SLE (96%) but have low diagnostic sensitivity (52%-70%) due to the brief time of presentation. The reasons for changes in anti-dsDNA antibody levels in vivo may be due to the following reasons: first, antibody concentrations vary with the timing of pathological changes in SLE, or non-nephropathic episodes are not always accompanied by anti-dsDNA antibodies; second, the antibodies may be deposited in renal tissues and undetectable in serum, or the detection techniques may not be sufficiently sensitive and specific; third, not all subclasses of anti-dsDNA antibodies are pathogenic and are associated with disease activity and nephropathy.

Alternative Names

double-stranded DNA IgG ELISA Kit

References

1. Yu H, et al. Clinical and Immunological Biomarkers for Systemic Lupus Erythematosus. Biomolecules. 2021 Jun 22;11(7):928.
2. Wang Y, et al. The Therapeutic Strategies for SLE by Targeting Anti-dsDNA Antibodies. Clin Rev Allergy Immunol. 2022 Oct;63(2):152-165.

Documents

pdf COA

pdf SDS

pdf Datasheet

QA & Reviews

Q & A

Ask a Question

Q: I am still currently looking for different dsDNA kits, and I am actually wondering from where the dsDNA was purified from in this kit? Or which type of donors were used?

A: The immobilized antigen is a highly purified preparation of plasmid dsDNA (>90% in supercoiled state), free from chromosomal DNA and protein (histones).

I hope this helps.

Customer Reviews