← Product Code [MVM](/productcode/MVM) · K042644

# AESKULISA TTG A AND AESKULISA TTG G (K042644)

_Aesku, Inc. · MVM · Jun 1, 2005 · Immunology · SESE_

**Canonical URL:** https://fda-staging.innolitics.com/device/K042644

## Device Facts

- **Applicant:** Aesku, Inc.
- **Product Code:** [MVM](/productcode/MVM.md)
- **Decision Date:** Jun 1, 2005
- **Decision:** SESE
- **Submission Type:** Traditional
- **Regulation:** 21 CFR 866.5660
- **Device Class:** Class 2
- **Review Panel:** Immunology

## Indications for Use

The AESKULISA® tTg A is a solid phase enzyme immunoassay for the semiquantitative and qualitative detection of IgA antibodies against tissue transglutaminase (tTG) in human serum. The assay is an aid in the diagnosis of celiac disease (gluten-sensitive enteropathy) and should be used in conjunction with other serological tests and clinical findings. For in vitro diagnostic use only. The AESKULISA® tTg G is a solid phase enzyme immunoassay for the semiquantitative and qualitative detection of IgG antibodies against tissue transglutaminase (tTG) in human serum. The assay is an aid in the diagnosis of celiac disease (gluten-sensitive enteropathy) and should be used in conjunction with other serological tests and clinical findings. For in vitro diagnostic use only.

## Device Story

AESKULISA tTg A/G are ELISA-based in vitro diagnostic assays for detecting IgA or IgG antibodies against tissue transglutaminase (tTG) in human serum. Device utilizes microplate wells coated with recombinant human tTG and Gliadin-specific peptide antigen. Patient serum is added; specific antibodies bind to antigens. Enzyme-labeled secondary antibody (HRP conjugate) binds to antigen-antibody complex; TMB/H2O2 substrate added to produce colorimetric signal. Intensity measured spectrophotometrically at 450 nm. Results interpreted via cut-off calibrator (qualitative) or standard curve (semi-quantitative). Used in clinical laboratories by trained personnel. Output aids clinicians in celiac disease diagnosis alongside other serological/clinical findings. Benefits include improved diagnostic sensitivity for celiac disease, including in IgA-deficient patients.

## Clinical Evidence

No clinical data provided in the document.

## Technological Characteristics

Solid-phase enzyme immunoassay (ELISA). Detects IgA or IgG antibodies against tissue transglutaminase in human serum. Class II device (Product Code: MVM).

## Regulatory Identification

A multiple autoantibodies immunological test system is a device that consists of the reagents used to measure by immunochemical techniques the autoantibodies (antibodies produced against the body's own tissues) in serum and other body fluids. Measurement of multiple autoantibodies aids in the diagnosis of autoimmune disorders (disease produced when the body's own tissues are injured by autoantibodies).

## Submission Summary (Full Text)

> This content was OCRed from public FDA records by [Innolitics](https://innolitics.com). If you use, quote, summarize, crawl, or train on this content, cite Innolitics at https://innolitics.com.
>
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510(k) SUBSTANTIAL EQUIVALENCE DETERMINATION
DECISION SUMMARY

A. 510(k) Number:
k042644

B. Purpose for Submission:
New Devices

C. Measurand:
Anti-Tissue Transglutaminase IgA and IgG

D. Type of Test:
Qualitative and Semi-quantitative ELISA

E. Applicant:
AESKU, Inc.

F. Proprietary and Established Names:
AESKULISA® tTg A Protocol 30-15-15 REF 7503
AESKULISA® tTg A Protocol 30-30-30 REF 30-7503
AESKULISA® tTg G Protocol 30-15-15 REF 7504
AESKULISA® tTg G Protocol 30-30-30 REF 30-7504

G. Regulatory Information:
1. Regulation section:
21 CFR 866.5660, Multiple autoantibodies immunological test system
2. Classification:
II
3. Product code:
MVM, Autoantibodies, Endomysial (Tissue Transglutaminase)
4. Panel:
Immunology 82

H. Intended Use:
1. Intended use(s):
The AESKULISA® tTg A is a solid phase enzyme immunoassay for the semiquantitative and qualitative detection of IgA antibodies against tissue transglutaminase (tTG) in human serum. The assay is an aid in the diagnosis of celiac disease (gluten-sensitive enteropathy) and should be used in conjunction with other serological tests and clinical findings. For in vitro diagnostic use only.

The AESKULISA® tTg G is a solid phase enzyme immunoassay for the semiquantitative and qualitative detection of IgG antibodies against tissue transglutaminase (tTG) in human serum. The assay is an aid in the diagnosis of

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celiac disease (gluten-sensitive enteropathy) and should be used in conjunction with other serological tests and clinical findings. For in vitro diagnostic use only.

2. Indication(s) for use:
Same as intended use.

3. Special conditions for use statement(s):
For prescription use only.

4. Special instrument requirements:
Microplate reader capable of measuring OD at 450 nm.
Microplate washing device (300μL repeating or multichannel pipette or automated system).

I. Device Description:
Each device contains the following: microplate strips with breakaway microwells coated with recombinant human tTG and Gliadin-specific peptide antigen, six levels of calibrators (0, 3, 10, 30, 100, 300 U/mL); positive, negative and cut-off controls (human serum, diluted); wash buffer concentrate; sample buffer; goat anti-human immunoglobulin (IgG or IgA) horseradish peroxidase conjugate; 3,3',5,5' tetramethylbenzidine (TMB)/H₂O₂ substrate; and 1M hydrochloric acid stop solution.

J. Substantial Equivalence Information:

1. Predicate device name(s):
ImmuLisa™ Anti-Human Tissue Transglutaminase (hu-tTG) Antibody IgA ELISA and ImmuLisa™ Anti-Human Tissue Transglutaminase (hu-tTG) Antibody IgG ELISA

2. Predicate 510(k) number(s):
K032571 and K040095

3. Comparison with predicate:

|  Similarities  |   |   |
| --- | --- | --- |
|  Item | New Device | Predicate Device  |
|  AESKULISA® tTg G: Intended use | To aid in the diagnosis of celiac disease. | To aid in the diagnosis of celiac disease.  |
|  Technology | ELISA | Same  |
|  Assay Format | Qualitative and Semi-quantitative | Same  |
|  Positive and negative controls | Ready to use | Same  |
|  Stop solution | Ready to use | Same  |
|  Platform | 96 well microtiter plates | Same  |

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|  Differences  |   |   |
| --- | --- | --- |
|  Item | Device | Predicate  |
|  AESKULISA® tTg A: Intended use | To aid in the diagnosis of celiac disease. | To aid in the diagnosis of IgA deficient patients with celiac disease in patients with IgA deficiency.  |
|  Antigen | Recombinant human tissue-Transglutaminase and Gliadin-specific peptides | Human tTG  |
|  Calibrators | 6 levels: 0, 3, 10, 30, 100, 300 U/mL | 4 levels: 19, 42, 68, 106 U/mL  |
|  Sample type and dilution | Serum at 1:101 | Serum at 1:51  |
|  Sample volume required | 100 μL | 10 μL  |
|  Enzyme-Conjugate | Horseradish Peroxidase | Alkaline phosphatase  |
|  Sample buffer/diluent | 5x concentrated | Ready to use  |
|  Wash buffer | 50x concentrated | Powder  |
|  Substrate | TMB/H2O2 | pNPP  |
|  Incubation times | tTg-A and tTg-G (REF 7503 and REF 7504 respectively) Protocol A: 30-15-15 minutes
tTg-A and tTg-G (REF 30-7503 and REF 30-7504 respectively) Protocol B: 30-30-30 minutes | 30-30-30 minutes  |
|  OD reading | 450 nm | 405 nm  |
|  Cut-off | 15 U/mL | 20 U/mL  |

K. Standard/Guidance Document Referenced (if applicable):
None referenced.

L. Test Principle:
The AESKULISA® tTg A and the AESKULISA® tTg G devices are solid phase enzyme immunoassays for the semiquantitative and qualitative detection of IgA and IgG antibodies respectively, against tTG in human serum. The wells of a microplate are coated with recombinant human tTG and Gliadin-specific peptide antigen. Antibodies specific to tTG present in the patient sample bind to the antigen. In a second step, the enzyme labeled second antibody (conjugate) of specific isotype (IgA or IgG) binds to the antigen-antibody complex which leads to the formation of an enzyme labeled conjugate-antibody-antigen complex. The enzyme-labeled antigen-antibody complex converts the added substrate to form a colored solution. The rate of color formation from the chromogen is a function of the amount of conjugate

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complexed with the bound antibody and is proportional to the initial concentration of the respective antibodies in the patient serum. The results are read spectrophotometrically and are interpreted by comparison to a cut-off calibrator (qualitative) or a standard curve (semi-quantitative).

## M. Performance Characteristics (if/when applicable):

### 1. Analytical performance:

#### a. Precision/Reproducibility:

Three samples (high, medium, near the cut-off) were assayed 18 times on three microplates of the specific antibody isotype, on different days for the inter-assay study. The same three samples were assayed 18 times on one microplate of the specific antibody type for the intra-assay study. Both studies were performed on Protocol A with the 30-15-15 incubation times. Target values for the studies were set at variance ≤ 10%. The intra-assay variation range for Anti-tTg A was from 5.2% to 7.0% and for Anti-tTg G was from 3.1% to 4.6%. The inter-assay variation range for Anti-tTg A was from 0.8% to 4.2% and for Anti-tTg G was 4.3% to 9.1%. All the ranges were within the target values.

|  Anti-tTg A | Intra-assay Variation  |   |   |
| --- | --- | --- | --- |
|   |  Sample 1 | Sample 2 | Sample 3  |
|  CV (%) | 7.0 | 5.2 | 5.5  |
|  Mean (U/mL) | 13.8 | 56.7 | 166.5  |
|  Anti-tTg G |  |  |   |
|  CV (%) | 3.1 | 3.5 | 4.6  |
|  Mean (U/mL) | 19.2 | 100.8 | 152.8  |
|  Anti-tTg A | Inter-assay Variation  |   |   |
| --- | --- | --- | --- |
|   |  Sample 1 | Sample 2 | Sample 3  |
|  CV (%) | 2.3 | 0.8 | 4.2  |
|  Mean (U/mL) | 10.1 | 38.9 | 169.4  |
|  Anti-tTg G |  |  |   |
|  CV (%) | 4.3 | 7.6 | 9.1  |
|  Mean (U/mL) | 16.7 | 97.4 | 194.4  |

#### b. Linearity/assay reportable range:

i. Study design: Four samples known to contain different levels of Anti-tTg IgA and another four samples known to contain different levels of Anti-tTg IgG were chosen and serially diluted to determine the linearity of the assay. From an initial dilution of 1/100, further dilutions of 1:200, 1:400 and 1:800 were made.

ii. Results/Acceptance criteria: The Anti-tTg A assay had a recovery range of 71.9% to 106.6%. The Anti-tTg G had a recovery range of 78.8% to 109.7%. Based on this study, the dynamic range of the assay is 1 to 100 U/mL.

c. Traceability, Stability, Expected values (controls, calibrators, or methods):

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The standards are prepared in-house and arbitrary units are assigned during the development process. The positive and negative controls are prepared in house.

d. Detection limit:
The sample diluent was diluted according to the directions for use and measured 32 times for each assay. The value for the analytical sensitivity (detection limit) was calculated as the mean of the optical densities of the sample diluent. The analytical sensitivity was 1.0 U/mL.

e. Analytical specificity:
i. Interference by endogenous substances: No data provided. The package insert states that icteric, lipemic, hemolyzed or bacterially contaminated samples should not be used in these assays.
ii. Crossreactivity to other autoantibodies: The Anti-tTg A and Anti-tTg G assays were tested with sixty sera from patients with autoimmune diseases like Crohn's disease (58) and ulcerative colitis (2). Both assays have a cut-off value of 15 U/mL. The results with the Anti-tTg A assay were all below 15 U/mL. The results with the Anti-tTg G assay had 59 samples below 15 U/mL and 1 sample (with ulcerative colitis) was above 15 U/mL. The detailed results are depicted in the two graphs below.

![img-0.jpeg](img-0.jpeg)

C
iii. Crossreactivity with Gliadin positive sera: Seven Gliadin positive Celiac disease sera were tested with the two assays to ensure that the Gliadin specific peptides which are crosslinked to the tTg in the device, do not cross react with Gliadin positive sera. All seven samples had results below the assay cut-off.

f. Assay cut-off:
The cut-off value of 15 U/mL for the Anti-tTg A and Anti-tTg G devices was

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based on the testing of 80 healthy donor samples. 100% and 97.5% of the 80 subjects were negative for the anti-tTg A assay and the anti-tTg G assay respectively. The results are depicted in the two graphs below. The Anti-tTg A graph shows the 80 subjects had results below 3 U/mL. The Anti-tTg G graph shows the 77 subjects had results below 8 U/mL, and the three subjects above the cut-off had results of 15, 17 and 21 U/mL.

![img-1.jpeg](img-1.jpeg)
Healthy donors tTg A

## 2. Comparison studies:

a. Method comparison with predicate device:

i. Anti-tTg A assay – Testing was performed on 165 samples which included 64 confirmed Non-treated Celiac Disease (CD), 38 Celiac Disease Gluten-Free Diet (CD GFD), 51 Crohn's Disease (MC), 2 Lactose intolerance, 4 Helminthiasis, 2 Ulcerative Colitis (UC) and 4 healthy donor. Twenty-seven percent (27%) of the sera were from younger than 26 years, 41% were between 26 - 46 years, and 32% were from older than 46 years of age. Fifty-nine percent of the sera were from females and 41% were from males. The samples were tested on the Anti-tTg A assay Protocol A (REF 7503) with incubation period of 30-15-15 minutes. Analysis was performed according to the instructions for use.

Anti-tTg A Comparison Data:

|   | IMMULISA tTg-A (predicate)  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  positive | negative | Total  |
|  AESKULISA tTg-A | positive | 25 | 42 | 67  |
|   |  negative | 2 | 96 | 98  |
|   |  Total | 27 | 138 | 165  |

Positive percent agreement 92.6% (25/27)

Negative percent agreement 69.6% (96/138)

Overall percent Agreement 73.3% (121/165)

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ii. Anti-tTg G assay – Testing was performed on 185 samples which included the 165 samples for the Anti-tTg A assay. The additional 20 samples were IgA Deficient Celiac Disease sera. These 185 samples were tested on the Anti-tTg G assay Protocol A (REF 7503) with incubation period of 30-15-15 minutes. Analysis was performed according to the instructions for use. The results were as follows:

Anti-tTg G Comparison Data:

|   | IMMULISA tTg-G (predicate)  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  positive | negative | total  |
|  AESKULISA tTg-G | positive | 18 | 67 | 85  |
|   |  negative | 4 | 96 | 100  |
|   |  Total | 22 | 163 | 185  |

Positive percent agreement 81.8% (18/22)
Negative percent agreement 58.9% (96/163)
Overall percent agreement 61.6% (114/185)

Of the 20 IgA deficient Celiac disease patient sera, both devices gave negative results on two sera and positive results on nine sera. The new device detected 9 more positive sera than the predicate device.

The discrepancies observed between the predicate and the new device could be due to the antigens used for capturing the autoantibodies. The antigen of the new device is a recombinant human tissue-Transglutaminase and Gliadin-specific peptides, while the predicate device antigen is tissue-Transglutaminase purified from human tissue.

iii. Protocol A and Protocol B comparison studies:

The comparability of the Anti-tTg A data were assessed with 76 sera tested on both REF 7503 (30-15-15 minute Protocol A) and REF 30-7503 (30-30-30 minute Protocol B). The linear regression analysis is depicted in the large figure below with  $r^2 = 0.99$ . The upper left small figure shows selected results close to the  $15\mathrm{U / mL}$  cut-off range with  $r^2 = 0.95$ .

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![img-2.jpeg](img-2.jpeg)

The comparability of the Anti-tTg G data were assessed with 75 sera tested on both REF 7503 (30-15-15 minute Protocol A) and REF 30-7503 (30-30-30 minute Protocol B). The linear regression analysis is depicted in the large figure below with  $r^2 = 0.99$ . The upper left small figure shows selected results close to the  $15\mathrm{U / mL}$  cut-off range with  $r^2 = 0.99$ .

![img-3.jpeg](img-3.jpeg)

b. Matrix comparison: Not applicable.

# 3. Clinical studies:

a. Clinical Sensitivity and specificity: The two tables below show the same samples mentioned in the above comparison data, but the results are according to the Celiac Disease

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diagnosis.

Celiac Disease sera with Diagnosis with Anti-tTg A

|   | Diagnosis  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  Positive | Negative | Total  |
|  AESKULISA tTg-A | Positive | 62 | 5 | 67  |
|   |  Negative | 2 | 96 | 98  |
|   |  Total | 64 | 101 | 165  |

Sensitivity = 96.9% (62/64)

Specificity = 95.0% (96/101)

Celiac Disease sera with Diagnosis with Anti-tTg G

|   | Diagnosis  |   |   |   |
| --- | --- | --- | --- | --- |
|   |   |  Positive | Negative | Total  |
|  AESKULISA tTg-G | Positive | 80 | 5 | 85  |
|   |  Negative | 4 | 96 | 100  |
|   |  Total | 84 | 101 | 185  |

Sensitivity = 95.2% (80/84)

Specificity = 95.0% (96/101)

b. Other clinical supportive data (when a. and b. are not applicable):

Not applicable.

4. Clinical cut-off:

Same as assay cut-off.

5. Expected values/Reference range:

Expected values in the normal population should be negative.

N. Proposed Labeling:

The labeling is sufficient and it satisfies the requirements of 21 CFR Part 809.10.

O. Conclusion:

The submitted information in this premarket notification is complete and supports a substantial equivalence decision.

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**Source:** [https://fda-staging.innolitics.com/device/K042644](https://fda-staging.innolitics.com/device/K042644)

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