The goal of this clinical trial is to evaluating the influence of single file rotary system for root canal instrumentation using different kinematics on postoperative pain, the primary purpose: measuring post-operative pain by Numerical rating scale in a cohort population treatyed in Jordan Universoit of science and technology post graduate clinics with previously intittaed root canal treatement and could include any of the following: sex/gender, age groups (18-65 ), and healthy volunteers.

The main questions it aims to answer [is/are:

• Is there any difference in postoperative pain after root canal treatment with different kinamtics
• Is this correlated with the level of cytokines measured taken from inside the canal system before and after treament whioch will be analyzed by the ELISA Kit for pain biomarkers.
• Is this related to in vitro debris extrusion, and can it validate previous research with the same methodology for debris extrusion measurement Researchers will compare 5 groups, which are Canal pro jeni, Continuous clockwise rotation (CCR), Optimum Torque Reverse (OTR), Forward reciprocating motion (REC), and Twisted Adaptive motion (TFA). All motions will be used with the One Curve file system Participants will report the level of pain after traatment level of cytokines will be measured by taking samples from the periapical area before, during, and after trematemtn](streamdown:incomplete-link)

This prospective, randomized, parallel-group clinical trial will evaluate whether the kinematics used during root canal instrumentation with a single-file system influence postoperative pain and the local expression of pain-related neuropeptides. The trial will be conducted in adult patients requiring endodontic treatment of mandibular molars. Eligible participants will be enrolled after clinical and radiographic assessment and after obtaining written informed consent.

Participants will be randomly allocated to one of five treatment groups. In all groups, canal shaping will be performed using the same One Curve nickel-titanium single-file system, while the kinematic motion used to activate the file will differ between groups. The tested kinematics will include CanalPro Jeni motion, continuous clockwise rotation, optimum torque reverse motion, forward reciprocating motion, and twisted adaptive motion. This design allows the effect of instrument motion to be assessed while maintaining the same file system across all study arms.

All treatments will be performed under standardized clinical conditions. Local anesthesia will be administered and the tooth will be isolated with a rubber dam. Access cavity preparation, canal identification, manual scouting, working length determination, and radiographic confirmation will be completed using a standardized approach. Initial irrigation will be performed before instrumentation, and the allocated kinematic protocol will then be used for canal preparation according to the study protocol.

Periapical fluid samples will be collected using sterile paper points at predefined procedural time points. The sampling schedule will include collection during the initial stage of canal preparation, after instrumentation, and at the follow-up visit. Samples will be transferred to sterile tubes and stored under appropriate laboratory conditions until analysis. The collected samples will be analyzed using enzyme-linked immunosorbent assay methods to quantify selected neuropeptides associated with inflammatory pain response, including substance P and calcitonin gene-related peptide.

The clinical treatment will be completed over two visits. At the first visit, the canals will be prepared using the allocated kinematic protocol and irrigated using a standardized irrigation sequence. Intracanal medicament will be placed between visits, and the tooth will be temporized. At the second visit, patient-reported pain and percussion sensitivity will be recorded, the temporary restoration and intracanal medicament will be removed, and periapical fluid will be collected before completion of the final irrigation protocol. The canals will then be dried, obturated using gutta-percha and resin-based sealer, and restored with an adhesive restorative material. A postoperative radiograph will be taken as part of routine treatment documentation.

Postoperative pain will be recorded by the participant at scheduled time points after treatment using a standardized pain scale. Clinical follow-up information and laboratory biomarker data will be collected using structured forms. The trial procedures will be performed by trained clinicians, and laboratory analysis will be performed according to the manufacturers' instructions for the selected assay kits. Data handling and storage will follow institutional requirements for clinical research.

The analysis will compare postoperative pain scores and biomarker levels between the five kinematic groups. Descriptive statistics will be used to summarize participant characteristics, pain scores, and biomarker concentrations. Appropriate statistical tests will be selected according to the type and distribution of the data. The analysis will assess whether differences in file kinematics are associated with differences in patient-reported postoperative pain and changes in pain-related neuropeptide levels.

Inclusion Criteria:

• Patients aged between 18-65 year
• Fully mature roots
• Asymptomatic
• Not taking neither antibiotics nor analgesics withing last 2 weeks
• ASA I and ASA II

Exclusion Criteria:

• patients that are younger or older than the selected age group 18-65 years - Immature roots
• Symptomatic, presence of sinus tract, fistula, or swelling.
• patient on medication antibiotics or analgesics withing last 2 weeks -Pregnant women,
• retreatment cases,
• allergic reactions
• resorption in the relevant tooth,
• periapical abscess,
• apical resorption, systemic diseases,ASA III-VI
• inability to understand the treatment procedure
• Any mesial canals with initial apical file larger than ISO 20

At the publication stage, the IPD will be shared

june 2026 to june 2027

biomarker analysis technician