Development of an In-House EphA2 ELISA for Human Serum and Measurement of Circulating Levels of EphA2 in Hypertensive Patients with Renal Dysfunction
Abstract
:1. Introduction
2. Materials and Methods
2.1. Monoclonal Antibodies to Human EphA2
2.2. Novel ELISA for Detecting of Human EphA2
2.3. Validation Procedures
2.3.1. Specificity
2.3.2. Accuracy
2.3.3. Lower Limit of Detection and Quantitation Limit
2.3.4. Precision
2.3.5. Measurement with a Commercially Available EphA2 ELISA
2.4. Patients and Sample Collection
2.5. Patients and Sample Collection
3. Results
3.1. Validation of the Novel In-House EphA2 ELISA
3.2. Comparison with the EphA2 ELISA Kit from Thermo Fisher Scientific
3.3. Circulating EphA2 Levels in Patients with Hypertension
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Characteristics | Total (n = 80) | eGFR (mL/min/1.73 m2) | p | |
---|---|---|---|---|
<60 (n = 64) | ≥60 (n = 16) | (<60 vs. ≥60) | ||
eGFR, mL/min/1.73 m2 | 39 (26, 56) | 35 (24, 43) | 71 (67, 79) | By design |
eGFR categories, no | ||||
G1+2 (≥60 mL/min/1.73 m2) | 16 | |||
G3a (45–59 mL/min/1.73 m2) | 14 | |||
G3b (30–44 mL/min/1.73 m2) | 26 | |||
G4 (15–29 mL/min/1.73 m2) | 24 | |||
Age, year | 72 ± 11 | 73 ± 11 | 68 ± 6 | 0.13 |
Male, no (%) | 52 (65.0) | 43 (67.2) | 9 (56.3) | 0.42 |
SBP, mmHg | 128 ± 13 | 128 ± 13 | 125 ± 11 | 0.42 |
DBP, mmHg | 73 ± 10 | 72 ± 10 | 78 ± 8 | 0.03 |
RAS blockers Tx, no (%) | 54 (67.5) | 43 (67.2) | 11 (68.8) | 0.91 |
UPCR, g/gCr | 0.38 (0.10, 1.00) | 0.50 (0.19, 1.46) | 0.09 (0.06, 0.26) | <0.0001 |
EphA2, pg/mL | 682 (434, 1006) | 775 (564, 1061) | 382 (329, 430) | <0.0001 |
Vendor | In-House | Thermo Fisher Scientific | ||
---|---|---|---|---|
EphA2 | r | p | r | p |
Age | 0.37 | 0.0007 | 0.03 | 0.82 |
SBP | 0.16 | 0.16 | 0.19 | 0.08 |
DBP | −0.26 | 0.02 | 0.07 | 0.57 |
eGFR | −0.83 | <0.0001 | −0.38 | 0.0004 |
UPCR | 0.61 | <0.0001 | 0.23 | 0.04 |
Variable | β | F | p |
---|---|---|---|
Age | 0.003 | 4.69 | 0.03 |
eGFR | −0.156 | 76.91 | <0.0001 |
UPCR | 0.031 | 3.33 | 0.07 |
DBP | 0.0003 | 0.03 | 0.86 |
R2 | 0.70 |
Baseline Characteristics (Units of Increase) | Model 1 | Model 2 | Model 3 | |||
---|---|---|---|---|---|---|
OR (95% CI) | p | OR (95% CI) | p | OR (95% CI) | p | |
Sex (female) | 0.96 (0.21, 4.50) | 0.96 | 1.02 (0.21, 4.92) | 0.98 | 0.82 (0.15, 4.50) | 0.82 |
Age (10 year) | 1.33 (0.50, 3.57) | 0.57 | 1.21 (0.40, 3.66) | 0.74 | 1.35 (0.50, 3.63) | 0.55 |
UPCR (1 quartile) | 3.79 (1.31, 10.99) | 0.014 | 3.98 (1.32, 12.02) | 0.01 | 3.81 (1.32, 11.00) | 0.01 |
EphA2 (1 quartile) | 4.74 (1.46, 15.42) | 0.0096 | 4.48 (1.36, 14.72) | 0.01 | 4.84 (1.47, 15.93) | 0.01 |
DBP (10 mmHg) | 0.74 (0.36, 1.53) | 0.42 | ||||
BP control (poor) | 1.58 (0.23, 11.06) | 0.64 |
Baseline Characteristics (Units of Increase) | Model 1 | Model 2 | Model 3 | |||
---|---|---|---|---|---|---|
OR (95% CI) | p | OR (95% CI) | p | OR (95% CI) | p | |
Sex (female) | 0.45 (0.08, 2.43) | 0.35 | 0.39 (0.07, 2.17) | 0.28 | 0.33 (0.06, 1.95) | 0.22 |
Age (10 year) | 0.74 (0.40, 1.38) | 0.45 | 0.68 (0.36, 1.30) | 0.24 | 0.88 (0.46, 1.66) | 0.68 |
UPCR (1 quartile) | 2.43 (1.09, 5.43) | 0.03 | 2.45 (1.07, 5.60) | 0.03 | 2.79 (1.16, 6.75) | 0.02 |
EphA2 (1 quartile) | 8.78 (2.68, 28.78) | 0.0003 | 10.42 (2.71, 40.07) | 0.0006 | 9.56 (2.81, 32.48) | 0.0003 |
DBP (10 mmHg) | 0.59 (0.24, 1.47) | 0.26 | ||||
BP control (poor) | 0.25 (0.03, 1.90) | 0.18 |
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Murakoshi, M.; Iwasawa, T.; Koshida, T.; Suzuki, Y.; Gohda, T.; Kato, K. Development of an In-House EphA2 ELISA for Human Serum and Measurement of Circulating Levels of EphA2 in Hypertensive Patients with Renal Dysfunction. Diagnostics 2022, 12, 3023. https://doi.org/10.3390/diagnostics12123023
Murakoshi M, Iwasawa T, Koshida T, Suzuki Y, Gohda T, Kato K. Development of an In-House EphA2 ELISA for Human Serum and Measurement of Circulating Levels of EphA2 in Hypertensive Patients with Renal Dysfunction. Diagnostics. 2022; 12(12):3023. https://doi.org/10.3390/diagnostics12123023
Chicago/Turabian StyleMurakoshi, Maki, Takumi Iwasawa, Takeo Koshida, Yusuke Suzuki, Tomohito Gohda, and Kazunori Kato. 2022. "Development of an In-House EphA2 ELISA for Human Serum and Measurement of Circulating Levels of EphA2 in Hypertensive Patients with Renal Dysfunction" Diagnostics 12, no. 12: 3023. https://doi.org/10.3390/diagnostics12123023