The majority of previous studies for early detection of Orthodontically Induced Inflammatory Root Resorption (OIIRR) were conducted to assess individual biomarkers which are representative of single biological pathway. To our Knowledge, no studies have been employed Gingival crevicular fluid (GCF) proteomic analysis to identify potential biomarkers for OIIRR. Therefore; the aim of this study is to use the advantage of testing a package of biomarkers via doing proteomic analysis of GCF which could help in making a more precise personalize diagnosis and treatments, thereby providing orthodontists additional information that cannot be deduced from clinical parameters.

Orthodontic patients with age range (18-years) indicated for bilateral maxillary first premolars extraction as part of their orthodontic treatment using specific fixed orthodontic appliance design with moderate anchorage were recruited for this study. Both of these premolars were allocated randomly to a heavy ( 225g) and light (25g) force in a split mouth design with a 1:1 allocation ratio. (GCF) was collected from the gingival sulcus of these teeth before extraction at six time intervals which are baseline, 1 hour, 1 day, 1 week , 2 weeks, 3 weeks and 4 weeks. Plaque and Gingival Indices were recorded for those patients at each timeline. Premolars will be extracted after 4weeks to be examined by stereomicroscopy for surface area evaluation of root resorption. Proteomic analysis will be carried out on the GCF sample. The proteomic analysis of the collected GCF will be obtained by using matrix-assisted laser desorption ionization/ time-of-flight mass spectrometry (MALDI-TOF/MS). MS spectra will be used to search human protein sequence data bases for protein identification and the development of "Protein Print" specific for these biological conditions.

Inclusion Criteria:

• Orthodontic patients indicated for extraction of bilateral upper first premolars,
• General healthy fit,
• Good oral hygiene with periodontal health maintenance (Plaque Index and Gingival Index ≤1),
• Lack of radiographic signs of idiopathic resorption,
• Radiographically both experimental teeth have two roots with approximately equal length and shape,
• The position of experimental teeth within the arch with no rotation.

Exclusion Criteria:

• Patients with impaction except impacted third molars for referral to do CBCT,
• Smokers,
• Patients on anti-inflammatory drugs at least three months before and during the study,
• History of oral habit,
• Previous orthodontic treatment,
• History of dental trauma,
• Previous dental treatment of the experimental teeth,
• Patient with heavy occlusion,
• Pathological lesion in the upper arch.

The GCF measures were coded by a clinician who bonded the appliance using patient ID number and the side of the arch (either right or left sides) to which orthodontic force (either heavy or light force) was allocated blindly for the investigator and outcomes assessor because the clinician could know the type of force from the shape of archwires. The surgeon who extract the teeth was also blinded. The extracted premolars were coded to be masked to the investigator who measured the root resorption areas. All outcome assessors of proteomic analysis as well as the statistician was blinded.