Bio-Functionalized Magnetic Nanoparticles for the Immunoassay of C-Reactive Protein and Procalcitonin in Cervicovaginal Secretions of Pregnant Women with Preterm Prelabor Rupture of Membranes to Predict Early-Onset Neonatal Sepsis

Figures & data

Figure 1 Depiction of magnetic nanoparticles. (A) A photograph of nanoparticles under dehydrated condition by scanning electron microscope (100,000X). (B) Illustration of a magnetic Fe₃O₄ nanoparticle coated with dextran and anti-CRP or ant-PCT antibodies.

Abbreviations: CRP, C-reactive protein; PCT, procalcitonin.

Figure 2 Illustration of the association between CRP and PCT and magnetic nanoparticles coated with anti-CRP or anti-PCT antibodies in the IMR assay. (A) Under the applied external alternating current magnetic fields, the magnetic nanoparticles oscillated and spun individually. (B) The magnetic nanoparticles became larger and clustered after binding with CRP or PCT, and thus oscillated and spun much slower than the original individual magnetic nanoparticles.

Abbreviations: CRP, C-reactive protein; PCT, procalcitonin; IMR, immunomagnetic reduction.

Figure 3 Magnitude and distribution of the anti-CRP and anti-PCT magnetic reagents determined by dynamic laser scattering. (A) The reagent with anti-CRP antibodies. (B) The reagent with anti-PCT antibodies.

Abbreviations: CRP, C-reactive protein; PCT, procalcitonin.

Table 1 The Mean Value, Standard Deviation and Coefficient of Variation of CRP and PCT

Biomarker	ϕ (ng/mL)	Mean (%)	SD (%)	CV (%)
CRP	0	0.90	0.052	5.71
	0.01	1.01	0.002	0.21
	0.1	1.05	0.003	0.34
	1	1.17	0.005	0.40
	10	1.37	0.004	0.29
	100	1.62	0.003	0.21
	300	1.71	0.004	0.23
	1000	1.70	0.006	0.33
PCT	0	0.89	0.055	6.14
	0.01	1.08	0.003	0.25
	0.1	1.14	0.004	0.32
	1	1.23	0.005	0.41
	10	1.37	0.006	0.42
	50	1.50	0.005	0.36
	100	1.57	0.005	0.30
	500	1.71	0.009	0.51

Abbreviations: CRP, C-reactive protein; PCT, procalcitonin; IMR, immunomagnetic reduction; SD, standard deviation; CV, coefficient of variation.

Figure 4 Calibration curves of (A) CRP and (B) PCT concentration-dependent IMR signals.

Abbreviations: CRP, C-reactive protein; PCT, procalcitonin.

Figure 5 Linearity and dynamic range of assaying (A) CRP and (B) PCT using the IMR assay.

Abbreviations: CRP, C-reactive protein; PCT, procalcitonin.

Figure 6 Individual relationships between detected biomarkers. (A) Cervicovaginal secretion PCT vs cervicovaginal secretion CRP, (B) cervicovaginal secretion CRP vs maternal serum WBC, (C) cervicovaginal secretion PCT vs maternal serum WBC, (D) maternal serum CRP vs cervicovaginal secretion CRP, and (E) maternal serum CRP vs maternal serum WBC.

Abbreviations: PCT, procalcitonin; CRP, C-reactive protein; WBC, white blood cell.

Table 2 Relationships Between Biomarkers and Chorioamnionitis

Biomarker	Without Chorioamnionitis (N = 24)	With Chorioamnionitis (N = 11)	P value
Cervicovaginal secretion CRP (ng/mL)	16.08 (10.32)	14.82 (20.45)	0.972
Cervicovaginal secretion PCT (ng/mL)	15.92 (31.88)	26.72 (61.88)	0.500
Maternal serum CRP (mg/L)	3.04 (4.02)	9.28 (7.62)	0.016*
Maternal serum WBC (/μL)	11,500 (5225)	13,200 (5300)	0.220

Notes: Continuous variables are analyzed by Mann–Whitney U-test and results are presented as median (interquartile range). *P < 0.05 was considered to be statistically significant.

Abbreviations: CRP, C-reactive protein; PCT, procalcitonin; WBC, white blood cell.

Table 3 Relationships Between Biomarkers and Early-Onset Neonatal Sepsis

Biomarker	Without Neonatal Sepsis (N = 18)	With Neonatal Sepsis (N = 15)	P value
Cervicovaginal secretion CRP (ng/mL)	15.19 (10.80)	20.11 (17.14)	0.262
Cervicovaginal secretion PCT (ng/mL)	9.54 (13.18)	45.99 (78.58)	0.039*
Maternal serum CRP (mg/L)	3.18 (3.53)	5.41 (9.38)	0.073
Maternal serum WBC (/μL)	12,250 (5300)	12,600 (4900)	0.704

Notes: Continuous variables are analyzed by Mann–Whitney U-test and results are presented as median (interquartile range). *P < 0.05 was considered to be statistically significant.

Abbreviations: CRP, C-reactive protein; PCT, procalcitonin; WBC, white blood cell.

Figure 7 Relationships between various biomarkers and early-onset neonatal sepsis. (A) Cervicovaginal secretion CRP, (B) cervicovaginal secretion PCT, (C) maternal serum CRP, and (D) maternal serum WBC.

Abbreviations: CRP, C-reactive protein; PCT, procalcitonin; WBC, white blood cell.

Figure 8 ROC curve of PCT for predicting early-onset neonatal sepsis using the IMR assay. The calculated cut-off PCT level was 20.60 ng/mL, and the area under the ROC curve was 0.71.

Abbreviations: ROC, receiver operating characteristic; PCT, procalcitonin.