Biotechnology Bulletin ›› 2021, Vol. 37 ›› Issue (5): 273-280.doi: 10.13560/j.cnki.biotech.bull.1985.2020-1181
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QU Huan1(), LI Cheng1, CHEN Rui1, LIAO Yi-jie1, CAO San-jie1,2,3, WEN Yi-ping1, YAN Qi-gui1, HUANG Xiao-bo1,2,3()
Received:
2020-09-18
Online:
2021-05-26
Published:
2021-06-11
Contact:
HUANG Xiao-bo
E-mail:1157759897@qq.com;rsghb110@126.com
QU Huan, LI Cheng, CHEN Rui, LIAO Yi-jie, CAO San-jie, WEN Yi-ping, YAN Qi-gui, HUANG Xiao-bo. Truncated Expression of the S1-CTD Fragment of Porcine Deltacoronavirus and Establishment of an Indirect ELISA for Detecting Its Antibody[J]. Biotechnology Bulletin, 2021, 37(5): 273-280.
Fig. 2 Identification of recombinant plasmid pET28a-S1CTD by PCR (A) and double digestion (B) A: Identification of recombinant plasmid pET28a-S1CTD by PCR (M: DL2000 DNA marker, 1: Target gene, N: Negative control, P: Positive control). B: Identification of recombinant plasmid pET28a-S1CTD by double digestion (M: DL5000 DNA marke, 1: Undigested recombinant plasmid, 2: Digested recombinant plasmid)
Fig. 3 The SDS-PAGE and Western blot analysis of recombinant S1-CTD protein A: The SDS-PAGE analysis of S1-CTD (M: Protein marker, 1: Post-induced BL21-pET28a, 2: Pre-induced pET28a-S1-CTD, 3: Post-induced pET28a-S1-CTD, 4: Hyperacoustic full bacteria after induction, 5: Hyperacoustic supernatant, 6: Hyperacoustic precipitation). B: Purification of S1-CTD protein (M: Protein marker, 1: Pre-lysing inclusion body protein, 2: Dissolved inclusion body protein, 3: Target protein eluted by 50 mmol/L imidazole, 4: Target protein eluted by 150 mmol/L imidazole, 5: Target protein eluted by 250 mmol/L imidazole). C: The western blot analysis of S1-CTD (M: Protein marker, 1-4: BL21-pET28a, pET28a-S1-CTD, pET-28a-E, pET-28a-N proteins reacted with pig anti-PDCoV positive serum, 5: pET28a-S1-CTD protein reacted with pig anti-PDCoV negative serum)
血清编号 Serum No. | 批内变异系数 Intra batch CV % | 批间变异系数 Inter batch CV % | ||
---|---|---|---|---|
±SD | 变异系数 CV/% | ±SD | 变异系数 CV/% | |
1 | 0.136±0.011 | 8.07 | 0.190±0.017 | 8.85 |
2 | 0.110±0.011 | 9.57 | 0.217±0.020 | 9.22 |
3 | 0.127±0.004 | 3.48 | 0.122±0.012 | 9.98 |
4 | 0.124±0.005 | 4.35 | 0.155±0.013 | 8.29 |
5 | 0.113±0.003 | 2.65 | 0.909±0.055 | 6.01 |
6 | 1.251±0.043 | 3.47 | 1.022±0.079 | 7.75 |
7 | 0.946±0.032 | 3.43 | 0.928±0.085 | 9.12 |
Table 1 Repeatability of the indirect ELISA
血清编号 Serum No. | 批内变异系数 Intra batch CV % | 批间变异系数 Inter batch CV % | ||
---|---|---|---|---|
±SD | 变异系数 CV/% | ±SD | 变异系数 CV/% | |
1 | 0.136±0.011 | 8.07 | 0.190±0.017 | 8.85 |
2 | 0.110±0.011 | 9.57 | 0.217±0.020 | 9.22 |
3 | 0.127±0.004 | 3.48 | 0.122±0.012 | 9.98 |
4 | 0.124±0.005 | 4.35 | 0.155±0.013 | 8.29 |
5 | 0.113±0.003 | 2.65 | 0.909±0.055 | 6.01 |
6 | 1.251±0.043 | 3.47 | 1.022±0.079 | 7.75 |
7 | 0.946±0.032 | 3.43 | 0.928±0.085 | 9.12 |
检测方法 Detection method | VNT | |||
---|---|---|---|---|
阳性数 Positive number | 阴性数 Negative number | 合计 Total | ||
阳性数 Positive number | 9 | 3 | 12 | |
阴性数 Negative number | 2 | 16 | 18 | |
间接ELISA | 合计 Total | 11 | 19 | 30 |
阳性符合率 Positive coincidence rate | 81.8% | |||
Indirect ELISA | 阴性符合率 Negative coincidence rate | 84.2% | ||
总符合率 Total coincidence rate | 83.3% |
Table 2 Comparative detection of serum samples by indirect ELISA and VNT
检测方法 Detection method | VNT | |||
---|---|---|---|---|
阳性数 Positive number | 阴性数 Negative number | 合计 Total | ||
阳性数 Positive number | 9 | 3 | 12 | |
阴性数 Negative number | 2 | 16 | 18 | |
间接ELISA | 合计 Total | 11 | 19 | 30 |
阳性符合率 Positive coincidence rate | 81.8% | |||
Indirect ELISA | 阴性符合率 Negative coincidence rate | 84.2% | ||
总符合率 Total coincidence rate | 83.3% |
来源 Origin | 样本数 Sample number | 阳性总数 Positive number | 阳性率 Positive rate/% |
---|---|---|---|
成都 Chengdu | 52 | 34 | 65.38 |
南充 Nanchong | 57 | 26 | 45.61 |
绵阳 Mianyang | 30 | 12 | 40 |
眉山 Meishan | 33 | 3 | 9.09 |
遂宁 Suining | 16 | 0 | 0 |
德阳 Deyang | 61 | 24 | 39.34 |
达州 Dazhou | 27 | 27 | 100 |
泸州 Luzhou | 9 | 8 | 88.89 |
宜宾 Yibin | 1 | 1 | 100 |
阿坝 Aba | 23 | 22 | 95.65 |
巴中 Bazhong | 90 | 84 | 93.33 |
凉山 Liangshan | 4 | 0 | 0 |
资阳 Ziyang | 1 | 0 | 0 |
雅安 Ya’an | 56 | 0 | 0 |
内江 Neijiang | 12 | 0 | 0 |
乐山 Leshan | 18 | 0 | 0 |
总数 Total | 490 | 241 | 49.18 |
Table 3 Clinical detection of PDCoV samples
来源 Origin | 样本数 Sample number | 阳性总数 Positive number | 阳性率 Positive rate/% |
---|---|---|---|
成都 Chengdu | 52 | 34 | 65.38 |
南充 Nanchong | 57 | 26 | 45.61 |
绵阳 Mianyang | 30 | 12 | 40 |
眉山 Meishan | 33 | 3 | 9.09 |
遂宁 Suining | 16 | 0 | 0 |
德阳 Deyang | 61 | 24 | 39.34 |
达州 Dazhou | 27 | 27 | 100 |
泸州 Luzhou | 9 | 8 | 88.89 |
宜宾 Yibin | 1 | 1 | 100 |
阿坝 Aba | 23 | 22 | 95.65 |
巴中 Bazhong | 90 | 84 | 93.33 |
凉山 Liangshan | 4 | 0 | 0 |
资阳 Ziyang | 1 | 0 | 0 |
雅安 Ya’an | 56 | 0 | 0 |
内江 Neijiang | 12 | 0 | 0 |
乐山 Leshan | 18 | 0 | 0 |
总数 Total | 490 | 241 | 49.18 |
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