Endometriosis and adenomyosis are two frequent estrogen-dependent gynecological diseases that lead to chronic pelvic pain, dysmenorrhea, abnormal uterine bleeding, and reproductive dysfunction.[1] Adenomyosis is characterized by the presence of endometrial glands and stroma within the myometrium, whereas endometriosis involves endometrium-like tissue outside the uterine cavity, resulting in chronic inflammation, fibrosis, and adhesions.[2] A recent systematic review and meta-analysis of the prevalence of focal adenomyosis and diffuse adenomyosis found pooled prevalence of 17% and 15%, respectively, with the global prevalence of adenomyosis in the general population estimated at 1% and of endometriosis at 5%.[3] The estimates are very widespread depending on the population studied and the diagnostic criteria employed.

Endometriosis is a significant global reproductive health problem, impacting 10% of women of reproductive age around the world.[4] It has been linked to chronic pelvic pain, intense dysmenorrhea, dyspareunia, and infertility, and is a detriment to physical, reproductive, and psychological health.[5] Traditionally, adenomyosis was believed to affect women in their mid to late reproductive years, but because of the advancement of imaging technology, more young women are being diagnosed with the condition.[6] Transvaginal ultrasonography has emerged as a valuable non-invasive diagnostic tool in both cases, enabling the identification of characteristic myometrial changes such as endometriomas and other disease features.[7] The symptoms of these various disorders may overlap, however, and there is a possibility that they may occur together.

These are both inflammatory and hormonal conditions that may affect circulating biochemical markers that are complex. Cancer antigen 125 (CA-125), a glycoprotein associated with epithelial tissues, has been investigated as a potential marker of endometriosis and adenomyosis.[8] It is seen to rise with peritoneal inflammation and irritation, but may be affected by disease extent and menstrual characteristics. A multicentre prospective study using 919 women assessed 54 blood-based biomarkers and found that CA-125 showed the best performance to identify endometriosis compared to the symptoms-only controls, with an area under the receiver operating characteristic curve (AUC) of 0.645 (95% CI: 0.600–0.690).[9] However, none of the assessed biomarkers proved to be sufficiently accurate to be used with confidence as a definitive test for diagnosis or exclusion.

C-reactive protein (CRP) is an acute phase protein produced by liver cells in response to inflammatory cytokines, and it may be a systemic inflammatory marker for chronic gynecological disorders.[10] Likewise, ferritin is mainly considered a marker of iron stores but is also an acute phase reactant and may be affected by chronic inflammation.[11] It is especially useful in women with heavy periods or if they lose blood for a long period. In 2024, a population-based prospective cohort study revealed that women with and without endometriosis experienced significantly different risk of iron deficiency (adjusted odds ratio: 1.46, 95% CI: 1.29–1.66).[12] These results emphasize the need to consider inflammatory and iron-related biochemical parameters as well as existing clinical and imaging data.

Despite these findings, evidence directly comparing CA-125, CRP, and ferritin between women with ultrasound-confirmed adenomyosis and endometriosis remains limited. The majority of the studies available have examined single biomarkers or changes in the affected women in comparison to either healthy or symptomatic women without a direct comparison between these two clinically overlapping diseases. Therefore, evaluating these three routinely available biochemical parameters together may provide a better understanding of the differences in their inflammatory and iron-related profiles and explore their potential value as adjuncts to clinical and ultrasonographic assessment. The present study aimed to compare serum levels of CA-125, C-reactive protein, and ferritin among women with ultrasound-confirmed adenomyosis and endometriosis, and to determine whether significant differences exist in these biochemical parameters between the two groups.

A comparative analytical cross-sectional study was conducted to compare serum CA-125, C-reactive protein (CRP), and ferritin levels among women with ultrasound-confirmed adenomyosis and endometriosis. The study was carried out in the Department of Obstetrics & Gynaecology, Abbasi Shaheed Hospital, Karachi, Pakistan. The biochemical tests were carried out in the Laboratory of the hospital according to standard laboratory procedures. The study lasted for 6 months from 1st December 2025 to 31st May 2026.

The sample size was determined using the Mean Difference calculator tool available on the OpenEpi version 3.01. Previous studies reported inconsistent estimates of mean and standard deviation for all three biomarkers in the two ultrasound-confirmed groups; therefore, a standardized effect size of 0.60 was assumed. With a 2-sided confidence level of 95%, power of 80%, and an equal allocation ratio of 1:1, a total of 44 participants were needed in each group, for a total of 88 participants. The final sample size was rounded to 100 women: 50 women with adenomyosis and 50 women with endometriosis, after adding about 10% to account for incomplete data or non-response.

A non-probability consecutive sampling technique was used.  The women were aged 18–45 years and presented with gynaecological complaints, with an ultrasound-confirmed diagnosis of either adenomyosis or endometriosis. Women who had characteristic ultrasonographic findings and were evaluated by an experienced radiologist or gynaecologist as having adenomyosis and/or endometriosis were eligible. Those who gave informed written consent were enrolled. To avoid confounding, women with co-occurring ultrasound-proven adenomyosis and endometriosis were excluded to allow for two comparison groups. Women with pregnancy, known gynecological malignancy, acute infectious conditions, chronic inflammatory or autoimmune disorders, significant hepatic or renal disease, or a history of recent blood transfusion were excluded. Those women who had received hormonal treatment for the past three months for either endometriosis or adenomyosis and those receiving iron supplements were not included, as these may affect the biochemical parameters studied.

After obtaining informed written consent, eligible participants were divided into two groups according to their ultrasonographic diagnosis. Group I consisted of 50 women having adenomyosis, and Group II consisted of 50 women having endometriosis. The data were collected using a structured data collection proforma; the data included the following information: age, BMI, parity, menstrual history, duration of symptoms, dysmenorrhea, chronic pelvic pain, abnormal uterine bleeding, heavy menstrual bleeding, infertility, and dyspareunia. Medical and gynecological histories were also taken.

The diagnosis was confirmed by ultrasonographic examination. The diagnosis of adenomyosis was made based on the classic features of myometrial cysts, asymmetrical myometrial thickening, fan-shaped shadowing, subendometrial lines and buds, and an irregular or interrupted junctional zone. Characteristic features, including deep endometriosis, ground-glass echogenicity, and the presence of ovarian endometrioma, were used to diagnose endometriosis. The diagnosis was made on the basis of ultrasound and clinical examination.

A 5 mL venous blood sample was drawn under aseptic conditions from each participant. The blood samples were sent to the lab of the institution for biochemical analysis. Serum CA-125 was assayed with a validated immunoassay, CRP was measured by an immunoturbidimetric method, and serum ferritin was measured by chemiluminescent immunoassay. All three investigations were conducted with standardised laboratory procedures, and the values were noted in U/mL for CA-125, mg/L for CRP, and ng/mL for ferritin. Measurement bias was minimized by blinding of laboratory personnel, when possible, to the diagnostic groups. All the demographic, clinical, ultrasonographic, and laboratory data were recorded in a standardized database for statistical analysis.

Data analysis was done through the use of Statistical Package for the Social Sciences (SPSS) version 26. The Shapiro–Wilk test was used to determine normality for quantitative variables such as age, BMI, duration of symptoms, serum CA-125, CRP, and ferritin levels. Values of normally distributed variables were presented as mean±SD, while the values of non-normally distributed variables were expressed as median and interquartile range. Qualitative variables such as parity, menstrual characteristics, and presenting symptoms were shown as frequencies and percentages.

An independent-samples t-test was used to compare normally distributed quantitative variables between the two groups, whereas the Mann–Whitney U test was applied for non-normally distributed variables. Categorical variables were compared by the chi-square test and Fisher's exact test. Receiver operating characteristic (ROC) curve analyses were conducted to evaluate the discriminatory ability of CA-125, CRP, and ferritin in distinguishing adenomyosis from endometriosis. The area under the curve (AUC), the 95% confidence intervals, the sensitivity, and specificity were computed, and the optimal cut-off was identified with the help of the Youden index. The independent association of the three biomarkers with the diagnosis of endometriosis was assessed using multivariable logistic regression analysis, when appropriate, after adjusting for the relevant confounding factors. A p-value of less than 0.05 was considered statistically significant.

The study included a total of 100 women, including 50 women diagnosed with ultrasound-verified adenomyosis and 50 women diagnosed with endometriosis. There were significant differences in the age and BMI of women with adenomyosis compared to women with endometriosis. The duration of symptoms was similar, but the extent of parity distribution varied considerably between the groups (Table 1).

In terms of clinical presentations, women with endometriosis were more likely to have chronic pelvic pain and dyspareunia, and women with adenomyosis were significantly more likely to have abnormal uterine bleeding and heavy menstrual bleeding. Dysmenorrhea was frequent in both groups, with no statistically significant difference (Table 2).

Characteristic disease-specific findings were identified by ultrasonographic assessment in both groups. Among the women who had adenomyosis, myometrial abnormalities such as asymmetrical thickening, myometrial cysts, and irregular junctional zones were noted. Among women with endometriosis, in contrast, ovarian endometriomas were found, as well as ground-glass features and signs of endometriosis in the depth (Table 3).

Comparison of biochemical parameters revealed significantly higher serum CA-125, CRP, and ferritin concentrations among women with endometriosis than among those with adenomyosis. All three biomarkers demonstrated statistically significant differences between the study groups (Table 4).

Categorical evaluation of elevated biomarkers also revealed a significantly greater proportion of elevated CA-125 and CRP in women with endometriosis. However, the frequency of elevated ferritin levels did not differ significantly between the two groups (Table 5).

ROC curve analysis demonstrated that CA-125 had the highest discriminatory performance for differentiating endometriosis from adenomyosis, followed by ferritin and CRP. All three biomarkers demonstrated statistically significant discriminatory ability, although their diagnostic performance was moderate (Table 6).

Table 1. Demographic and baseline characteristics of study participants (n = 100)

Table 2. Clinical characteristics of the study participants

Table 3. Ultrasonographic characteristics of women with adenomyosis and endometriosis

Table 4. Comparison of serum CA-125, CRP, and ferritin levels between the two groups

Table 5. Comparison of elevated biochemical markers between the two study groups

Table 6. ROC curve analysis of serum biomarkers for differentiating endometriosis from adenomyosis

The present study compared serum CA-125, C-reactive protein (CRP), and ferritin levels among 100 women with ultrasound-confirmed adenomyosis and endometriosis. The results showed significantly elevated levels of all three biomarkers in women with endometriosis, with CA-125 having the highest discriminatory power, followed by ferritin, and then CRP. These findings provide an opportunity to explore the differences in biochemical profiles between two gynecological conditions with overlapping clinical manifestations.

The present study revealed significant differences between women with adenomyosis and women with endometriosis with respect to age, BMI, and parity. This result was consistent with the clinical difference reported for these conditions. Tang et al. (2023) also showed a relationship between patient characteristics, disease subtype, and biomarker concentration in their study of CA-125 in adenomyosis.[13] Likewise, Burghaus et al. (2023) reported on the multicentre study of 919 women, pointing to the difficulties in differentiating adenomyosis from endometriosis.[14] These results highlight the need to take into account demographic and clinical factors when assessing biochemical differences between the two groups.

significantly higher in women with endometriosis than in those with adenomyosis in the present study. It had moderate discriminatory ability with an AUC of 0.742 in its ROC analysis. This result was similar to a multicentre study by Burghaus et al. (2023) that found CA-125 had an AUC of 0.645 when it came to the identification of endometriosis among women with symptoms.[14] However, Tang et al. (2023) found that the level of CA-125 was associated with the severity of the disease, and the optimum cut-off value for dysmenorrhea was determined to be 35 U/mL for CA-125.[13]

 In another retrospective study of 200 women with adenomyosis, Yang et al. (2024) found significantly elevated CA-125 levels in their group compared to controls.[15] Overall, all of these studies suggest a link between CA-125 and both conditions, but the discriminatory ability of CA-125 may differ depending on the comparison groups and the characteristics of the condition.

In the present study, CRP was also significantly elevated in the women who had endometriosis, with less discriminatory power than CA-125. In a case-control study of 59 women with histologically proven endometriosis and 28 controls, Kasoha et al. (2025) reported no difference in CRP levels between the groups.[16] In another study, a prospective study of 103 women with endometriosis and 103 healthy women, Sarikaya Kurt et al. (2025) found that there were lower levels of CRP in the endometriosis group than in healthy controls. The results contrast with the current study and indicate that the levels of circulating CRP do not always reflect the inflammatory activity of endometriosis. These disparities may be attributable to variations in study populations, the types of controls, or inflammatory disease.[17]

In the current study, the concentration of ferritin was significantly higher in women with endometriosis when compared to women with adenomyosis. However, previous research has demonstrated a considerable association between endometriosis and iron deficiency. In a population-based prospective cohort study, Gete et al. (2024) found that women with endometriosis were 46% more likely to have iron deficiency than women without the condition.[18] In a multicentre cross-sectional study of 251 women, Goldberg et al. (2025) found that 53.4% of women had been diagnosed with iron deficiency. These results were not directly relevant to our group with endometriosis and their higher levels of ferritin. Ferritin is also an acute phase reactant and may be affected by iron status and inflammation.[19] Thus, any difference observed should be interpreted with caution and not necessarily as a sign of increased iron stores or availability.

In the present study, ferritin also showed good discrimination, with an AUC of 0.714. Huang et al. (2024), while investigating serum CA-125 and prostaglandin E2 in women with adenomyosis, reported changes in CA-125 following treatment, supporting its potential relationship with disease activity.[20] In another study, Schoeman et al. (2025) examined plasma protein biomarkers in 805 participants and found a panel of 10 proteins that had diagnostic potential for endometriosis. Their study, however, showed the continued interest in non-invasive biochemical disease assessment methods, albeit different from the one used by them.[21]

In general, ROC analysis showed that CA-125 was the most useful single marker, followed by ferritin and CRP. However, the AUC values suggested a moderate discriminatory ability. Results from Burghaus et al. (2023) also showed that none of the blood biomarkers was accurate enough for a standalone diagnosis of or exclusion of endometriosis or adenomyosis.[14] Hence, these biochemical parameters can be used as supplementary to clinical and ultrasonographic evaluation and not as an alternative to the existing routine diagnostic techniques.

LIMITATIONS

There were some limitations in the study. Its cross-sectional design did not allow assessment of temporal relationships between biomarker levels and disease progression. The results might only apply to the relatively small sample size and single center. In addition, there may have been some misclassification in diagnosis based on ultrasound rather than surgery or histopathology. Menstrual cycle phase, hormonal changes, iron status, and subclinical inflammatory states may also have affected the levels of serum biomarkers. This study did not measure changes in the biomarkers over time or after treatment. These findings should be confirmed in larger multicentre studies with uniform diagnostic and laboratory procedures.

Serum CA-125, CRP and ferritin levels were significantly higher among women with endometriosis than among those with adenomyosis in the present study. CA-125 demonstrated the highest discriminatory performance, followed by ferritin and CRP. However, the moderate diagnostic accuracy of these biomarkers indicates that none should be considered an independent diagnostic tool. Their evaluation may add to clinical examination and ultrasonography as a means of biochemical information. Larger population studies and modified analyses are needed to determine their clinical usefulness.

Martire, F.G., et al., Endometriosis and adenomyosis: from pathogenesis to follow-up. Current issues in molecular biology, 2025. 47(5): p. 298.

2. Khan, K.N., A. Fujishita, and T. Mori, Pathogenesis of human adenomyosis: current understanding and its association with infertility. Journal of Clinical Medicine, 2022. 11(14): p. 4057.
3. Wang, M.-H., et al., Global prevalence of adenomyosis and endometriosis: a systematic review and meta-analysis. Reproductive Biology and Endocrinology, 2025. 23(1): p. 148.
4. Xu, S., et al., Global, regional, and national burden of endometriosis among women of childbearing age from 1990 to 2021: a cross-sectional analysis from the 2021 global burden of disease study. International Journal of Surgery (London, England), 2025. 111(9): p. 5927.
5. Della Corte, L., et al., The burden of endometriosis on women’s lifespan: a narrative overview on quality of life and psychosocial wellbeing. International journal of environmental research and public health, 2020. 17(13): p. 4683.
6. Protopapas, A., et al., Adenomyosis: Disease, uterine aging process leading to symptoms, or both? Facts, views & vision in ObGyn, 2020. 12(2): p. 91.
7. Daniilidis, A., et al., Transvaginal ultrasound in the diagnosis and assessment of endometriosis—an overview: how, why, and when. Diagnostics, 2022. 12(12): p. 2912.
8. Feduniw, S., et al., Current evidence on CA-125 levels in differentiation between endometriomas and endometriosis-associated ovarian cancer–A systematic review and meta-analysis. Journal of Endometriosis and Pelvic Pain Disorders, 2024. 16(3-4): p. 154-159.
9. Burghaus, S., et al., Multicenter evaluation of blood‐based biomarkers for the detection of endometriosis and adenomyosis: A prospective non‐interventional study. International Journal of Gynecology & Obstetrics, 2024. 164(1): p. 305-314.
10. Putri, N.R., M.H. Hutabarat, and I.M.A. Subadiyasa, Role of C-Reactive Protein as an Inflammation Biomarker in Women of Reproductive Age: Literature Review. International Journal of Medicine and Public Health, 2025. 2(1): p. 76-86.
11. Fonseca, Ó., et al., New perspectives on circulating ferritin: its role in health and disease. Molecules, 2023. 28(23): p. 7707.
12. Gete, D.G., et al., Risk of Iron Deficiency in Women With Endometriosis: A Population-Based Prospective Cohort Study. Women's Health Issues, 2024. 34(3): p. 317-324.
13. Tang, Y., et al., Serum CA125 as a biomarker for dysmenorrhea in adenomyosis. Int J Gynaecol Obstet, 2023. 163(1): p. 131-139.
14. Burghaus, S., et al., Multicenter evaluation of blood-based biomarkers for the detection of endometriosis and adenomyosis: A prospective non-interventional study. Int J Gynaecol Obstet, 2024. 164(1): p. 305-314.
15. Yang, F., et al., CA125-associated activated partial thromboplastin time and thrombin time decrease in patients with adenomyosis. Journal of Multidisciplinary Healthcare, 2024: p. 251-261.
16. Kasoha, M., et al., Evaluation of inflammatory serum parameters as a diagnostic tool in patients with endometriosis: a case-control study. Sci Rep, 2025. 15(1): p. 20172.
17. Kurt, D.S., et al., Evaluation of serum-based inflammatory and haematological markers in patients with endometriosis: A case–control study. European Journal of Obstetrics & Gynecology and Reproductive Biology, 2025: p. 114751.
18. Gete, P.D.G., et al., Risk of iron deficiency in women with endometriosis: a population-based prospective cohort study. Women's Health Issues, 2024. 34(3): p. 317-324.
19. Goldberg, H.R., et al., High prevalence of undiagnosed iron deficiency in endometriosis patients: A cross‐sectional study. International Journal of Gynecology & Obstetrics, 2025. 168(3): p. 1321-1327.
20. Huang, Y. et al., Changes in serum CA125 and PGE2 before and after high-intensity focused ultrasound combined with GnRH-a in treatment of patients with adenomyosis. Open Medicine, 2024. 19(1): p. 20230794.
21. Schoeman, E., et al., Identification of plasma protein biomarkers for endometriosis and the development of statistical models for disease diagnosis. Human Reproduction, 2025. 40(2): p. 270-279.