Abstract Introduction For women experiencing bothersome genital arousal, pain, or other dysesthesia symptoms (e.g. feelings of itching, burning, pulsating, tearing), it is important to ascertain whether symptoms are secondary to pathology in the genital area, or are referred from upstream pathology in the pelvis or perineum, cauda equina, spine, and/or brain regions. Specific understanding of the location of the trigger associated with the genital dysesthesia is critical to directing treatments for disease modification management. Objectives The aim of this study is to review the methodology of minimally-invasive genital anesthesia testing including the clitoral anesthesia test (CAT – Figures 1a, 1b), vestibular anesthesia test (VAT – Figure 2), urethral anesthesia test (UAT) and/or Bartholin cyst anesthesia test (BAT). Methods Prior to genital anesthesia testing the patient documents the symptom score intensity on a scale of 0 to 10 where 10 is most severe. It is critical that the patient have significant symptoms in the specific genital region(s) (clitoris, vestibule, urethral meatus and/or Bartholin’s cyst); the patient may need to perform some maneuvers to trigger the dysesthesia symptoms and/or reduce or discontinue medications that offer relief from the dysesthesia. The area to be tested is demarcated under vulvoscopic examination with a marking pen. Using a cotton swab as an applicator, benzocaine (20%), lidocaine (8%), tetracaine (8%), (BLT) is first applied to the inner arm to assess for allergies. Assuming there is no allergic reaction, the BLT is applied generously to the specific genital region. If needed, local subcutaneous administration of 1% lidocaine using a 30-gauge needle can supplement the anesthesia. After 10 minutes, provocative maneuvers are performed to determine whether or not there is clinically significant symptom reduction from the pre-genital anesthesia test symptom score. Results A positive genital anesthesia test is consistent with clinically significant symptom reduction from the pre-genital anesthesia test symptom score intensity. A positive genital anesthesia test implies that the local genital region (clitoris, vestibule, urethral meatus and/or Bartholin’s cyst) is the trigger of the genital dysesthesia so appropriate treatment strategies may be offered. For example, a positive CAT may require lysis of adhesions. A positive VAT may require hormone management for hormone mediated vestibulodynia, or vestibulectomy for neuro-proliferative vestibulodynia. A positive UAT may require surgical reconstruction of the urethral meatus. A positive BAT may require marsupialization. A negative genital anesthesia test is consistent with upstream pathology leading to testing of the integrity of the pudendal nerve or assessment of the cauda equina with neuro-genital testing and lumbar/sacral MRI. Conclusions Genital anesthesia testing is an excellent minimally invasive, in office diagnostic test to help localize the trigger of the genital arousal, pain, or dysesthesia in the genital region. Recognizing the location of the trigger associated with the genital dysesthesia as emanating from the genital region rather than the pelvis or perineum, cauda equina, spine, and/or brain region, will allow for disease modification and specific management strategies of the clitoris, vestibule, urethral meatus and/or Bartholin cyst. Disclosure No.