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Table 1 Characteristics of samples, method of collection and processing, biomarker detection technique, radiographic method used, and conclusions

From: Non-invasive methods for the assessment of biomarkers and their correlation with radiographic maturity indicators — a scoping review

  Study Type of study Sample Age group Gender Type of sample Method of sample collection Parameters assessed Sample processing Biomarker detection technique Radiographic method used Conclusion
Saliva
 IGF1 Nayak et al. [23] Cross-sectional 45 subjects 7 to 23 years 24 females, 21 males Unstimulated parotid saliva Saliva was collected with a modified Lashley cup for a duration of 30 min. a. Salivary flow rate (ml/min)
b. IGF1 levels(ng/ml)
c. IGF1 secretion rate (ng/min)
Centrifugation for 10 min at 1500×g at room temperature, and stored at – 20 °C until analysis. Immunoradiometric assay (IRMA) Lateral cephalogram—Quantitative Cervical Maturation System (QCVM) by Chen et al. (2008) Results showed that the IGF1 levels in saliva and the rate of secretion were lower in QCVM I followed by an increase in QCVM II and a decrease in QCVM III and IV.
 BALP Wijaya et al. [24] Cross-sectional 136 subjects 8 to 18 years 64 males, 72 females Unstimulated whole saliva at 9 am Passive drooling method, after distilled water mouth rinse a. Total protein content
b. BALP levels(pg/ml)
Samples were centrifuged and stored in the icebox until analysis. Enzyme-linked immunosorbent assay (ELISA) Lateral cephalogram—Cervical Vertebral Maturation Index by Baccetti et al. [10] No significant differences were found between the groups.
Hegde et al. [25] Cross-sectional 90 subjects 6 to 19 years Not specified Unstimulated whole saliva Passive drooling method a. Salivary BALP levels(U/I) Not specified. Enzyme-linked immunosorbent assay (ELISA) Hand-wrist radiographs—Hagg and Taranger They concluded that the BALP levels in saliva increased in peak pubertal stage and showed a gradual increase from Subgroup S0 to Subgroup MP3.
 ALP Alhazmi et al. [26] Cross-sectional 79 subjects 7 to 23 years 48 females, 31 males Unstimulated whole saliva was collected between 9 am to 12 pm Passive drooling method for a period of 5 min a. Salivary ALP activity (mU/mg)
b. Total protein concentration (mg/mL)
Samples were centrifuged and stored at – 80 °C until analysis. Colorimetric assay Lateral cephalograms—Cervical Vertebral Maturation Index by Baccetti et al. [10] - Protein concentration was higher in CVMS III and V.
- Higher ALP levels in saliva were found in CVMS I.
The salivary activity of ALP was higher in males, suggesting their increased growth potential and longer growth spurt duration than in females.
Tarvade et al. [27] Cross-sectional 120 subjects 10 to 15 years 60 males, 60 females Unstimulated whole saliva Not specified a. Salivary ALP levels (IU/L) Samples were centrifuged and stored in an icebox before analysis. Colorimetric assay Middle phalanx third finger—MP3 staging by Hagg and Taranger The levels of salivary ALP reached a peak in the G stage of MP3 followed by a decrease in the H stage. They also found that both boys and girls showed ALP levels correlating well with the G stage of MP3
Irham et al. [28] Cross-sectional 57 subjects 8 to 15 years 57 females Unstimulated whole saliva was collected between 10 am to 12 pm Not specified a. Salivary ALP levels (IU/L) Samples were centrifuged for 2 min for 10,000 rpm and stored at – 80 °C. Colorimetric assay Lateral cephalogram—Cervical Vertebral Maturation Index by Hassel and Farman Increased activity of salivary ALP was seen in the pubertal phase. The difference between ALP levels of pre-pubertal and pubertal, and pubertal and post-pubertal was statistically significant.
 VEGF Sharmila et al. [29] Cross-sectional 90 subjects 6 to 20 years Not specified Unstimulated whole saliva The subjects were asked to expectorate saliva in a 15-ml Tarzon centrifuge tube after retaining it in the mouth for 5 min. a. Salivary IGF1 levels (ng/ml)
b. Salivary VEGF levels (pg/ml)
Centrifugation for 10 min at 1500×g at room temperature and stored at – 20 °C until analysis. Enzyme-linked immunosorbent assay (ELISA) Lateral cephalogram—Cervical Vertebral Maturation Index by Hassel and Farman No statistical difference was found among the pre-pubertal, pubertal, and post-pubertal stages for VEGF. But IGF1 levels were found to increase in the pubertal growth phase.
 DHEA Sangeeth et al. [30] Cross-sectional 66 subjects 9 to 18 years 33 males, 33 females Unstimulated whole saliva was collected at 10 am Passive drooling in a plastic vial a. Salivary DHEA levels (pg/ml) Centrifugation at 3000 rpm for 15 min. The samples were stored at – 4 °C immediately after collection (within 30 min) and transferred to – 20 °C within 4 h of collection. DHEA Immuno Assay Lateral cephalogram—Cervical Vertebral Maturation Index by Baccetti et al. [10] They found a gradual increase in DHEA concentration in saliva from stage 1 to stage 6 with the highest levels found in stage 5 and 6.
GCF
 ALP Perinetti et al. [31] Cross-sectional 72 subjects 7.8–17.7 years 45 females, 27 males GCF collected using #25 standardized sterile paper strips inserted 1 mm into the gingival crevice and left in situ for 60 s Two sites on each maxillary and mandibular central incisor a. GCF ALP activity (mU/sample) Samples transferred to plastic vials and immediate storage at – 80 °C until analysis. The total ALP activity was determined by monitoring the increase in spectrophotometric absorption, at 405 nm. The absorbance was converted into enzyme activity units (1 unit 5 1 mmol of p-nitrophenol released per minute at 37°C) and expressed as total activity in mU per sample. Lateral cephalogram—Cervical Vertebral Maturation Index by Baccetti et al. [10] A twofold increase in GCF ALP activity was found during the pubertal phase than during the pre-pubertal and post-pubertal phases.
Perinetti et al. [32] Cross-sectional 50 subjects 7.8–17.7 years 31 females, 19 males GCF collected using #25 standardized sterile paper strips inserted 1 mm into the gingival crevice and left in situ for 60 s Two sites on each maxillary and mandibular central incisor a. Total protein content (μg/sample)
b. Total GCF ALP activity (mU/sample)
c. Normalized GCF ALP activity (mU/μg proteins)
Four samples were pooled and immediately stored at – 80 °C The total ALP activity was determined by monitoring the increase in spectrophotometric absorption, at 405 nm. The absorbance was converted into enzyme activity units (1 unit 5 1 mmol of p-nitrophenol released per minute at 37°C) and expressed as total activity in mU per sample. Lateral cephalogram—Cervical Vertebral Maturation Index by Baccetti et al. [10] The total activity of GCF ALP was found to be significantly higher in the pubertal growth phase than in the pre-pubertal and post-pubertal growth phases in both the maxillary and mandibular sites.
 VIT DBP Wen et al. [33] Cross-sectional 40 subjects 6.8–24 years 20 males, 20 females GCF was collected between 8 am and 10 am. Paper strips were inserted and left in situ for 30 s. From mesiolabial and distolabial sites of upper central incisors a. Vitamin D binding protein (μg)
b. Serotransferrin (μg)
50 μl phosphate-buffered solution was added into the Eppendorf tubes containing four paper points and shaken at 4 °C for 10 min and then centrifuged at 9000 rpm for 5 min. Enzyme-linked immunosorbent assay (ELISA) Cervical vertebral maturation method proposed by Franchi et al. [35] GCF levels of vitamin DBP and Serotransferrin were found to be significantly greater in pubertal compared to the post-pubertal growth phase.
Urine
 IGF1 Sinha et al. [34] Cross-sectional 72 subjects 8–20 years 72 females Random morning midstream urine samples were collected   a. Urine IGF1 levels (ng/ml) The urine samples were pipetted in Eppendorf tubes and ultracentrifuged and stored at – 80 °C Enzyme-linked immunosorbent assay (ELISA) Lateral cephalogram—Cervical Vertebral Maturation Index by Hassel and Farman Increased urinary IGF1 levels were found in CVMI stage 4 corresponding to the mean age of 13.67 years.