# Canine Pregnancy Diagnosis: Ultrasound and Hormonal Assays


## Key Takeaways

- **Ultrasonography** is the most versatile diagnostic tool, capable of visualizing gestational sacs from approximately day 20-22 post-ovulation and fetal heartbeat by day 25-28, allowing for viability assessment and moderate fetal count accuracy, particularly between days 28-35.
- **Relaxin assays** are pregnancy-specific hormonal tests, with serum relaxin becoming detectable from approximately day 22-25 post-ovulation, offering reliable confirmation from day 28 onwards, but do not assess fetal viability or count.
- **Progesterone assays** are not diagnostic for pregnancy itself but are crucial for confirming ovulation and assessing corpus luteum function, with a typical threshold of >2 ng/mL indicating luteal activity, essential for timing other diagnostic tests.
- **Timing is critical** for both methods; ultrasonography before day 20 or relaxin testing before day 22 carries a significant risk of false negatives, necessitating repeat testing or later examination for accurate diagnosis.
- **Discordant results** between ultrasound and relaxin assays, particularly a positive relaxin with no ultrasound findings, warrant repeat testing and consideration of early resorption or assay error, with ultrasound findings of fetal heartbeat being the most definitive indicator of viability.
- **Fetal counting accuracy** via ultrasound is moderate and improves after day 30 but is prone to error due to fetal overlap; radiography after day 45 offers a more reliable count for litter size estimation in late gestation.

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Confirming pregnancy in the bitch is a core theriogenology service that shapes breeding management, prenatal care planning, and owner expectations. This article compares the principal diagnostic methods, ultrasonography and hormonal assays, with attention to timing, accuracy, and interpretation. It serves the practicing veterinarian who must select a test, schedule it correctly relative to ovulation or breeding, and interpret results that may be ambiguous. The focus is diagnostic reasoning only, management of the pregnant bitch and whelping are outside this article's scope.

The bitch presents a unique diagnostic challenge because her gestational period is short and ovulation timing is frequently unknown. Breeding dates alone do not provide due dates with adequate accuracy, which complicates both pregnancy diagnosis and subsequent gestational age estimation [Lopate's review of gestational age assessment](https://pubmed.ncbi.nlm.nih.gov/18534674/). A negative result on any single test performed too early does not exclude pregnancy, and a positive result on a hormonal assay must be weighed against the possibility of luteal pathology. The clinician therefore needs a clear framework for when each test is informative and what its failure modes are.

## At a Glance

| Parameter | Ultrasound | Relaxin assay | Progesterone assay |
|---|---|---|---|
| Earliest positive result | Gestational sacs visible from approximately day 20 to 22 after ovulation | Serum relaxin detectable from approximately day 22 to 25 after ovulation | Not diagnostic for pregnancy, reflects luteal function only |
| Optimal window | Day 25 to 35 for confirmation, later for fetal count and viability | Day 28 or later for reliable confirmation | Day 20 to 25 after ovulation to assess luteal sufficiency |
| What it confirms | Presence of gestational sacs, embryos, fetal heartbeat | Presence of placental relaxin-secreting tissue | Ovulation occurred and corpus luteum is functional |
| Fetal viability assessment | Yes, via heartbeat and fetal movement | No | No |
| Fetal count accuracy | Moderate, improves after day 30 but litter size estimates carry error | Not possible | Not possible |
| Principal false negative risk | Examination before day 20 or operator inexperience | Testing before day 22 | Not applicable |
| Principal false positive risk | Mistaking uterine fluid or cysts for gestational sacs | Rare, persistent luteal tissue can produce low levels | Not applicable |

## Reproductive Physiology Relevant to Diagnosis

The canine estrous cycle dictates when each diagnostic method becomes valid. Ovulation occurs approximately 48 hours after the luteinizing hormone surge, and the oocytes require an additional 48 to 72 hours to mature before fertilization is possible. The corpus luteum forms rapidly and secretes progesterone throughout gestation, with luteal function maintained by pituitary luteinizing hormone instead of placental luteotropin. This physiology has a direct diagnostic consequence: progesterone is present in high concentrations in both pregnant and non-pregnant bitches during diestrus, so a progesterone measurement can confirm ovulation but cannot distinguish pregnancy from a prolonged luteal phase.

Relaxin is the only practical pregnancy-specific hormone in the bitch. It is produced by the developing placenta, specifically the trophoblast, and enters the maternal circulation in detectable quantities only when functional placental tissue exists. The Society for Theriogenology maintains professional resources on reproductive physiology and diagnostic protocols that the clinician can consult for detailed normative data [Society for Theriogenology resources](https://www.therio.org/). Because relaxin is placental in origin, its detection is strong evidence of pregnancy, although the assay's sensitivity depends on the gestational stage at sampling.

## Ultrasonography: Technique and Timing

Transabdominal ultrasonography is the most versatile diagnostic tool for canine pregnancy. A high-frequency linear or microconvex transducer, typically 7.5 to 10 MHz, provides the resolution needed to identify small gestational structures in the first half of gestation. The bitch is examined in dorsal or lateral recumbency, and the urinary bladder serves as an acoustic window to the uterine body and horns. Hair should be clipped or parted and coupling gel applied generously to minimize artifact.

The earliest reliable ultrasound finding is the gestational sac, an anechoic fluid-filled structure within the uterine horn, visible from approximately day 20 to 22 after ovulation. The embryonic vesicle appears as a small echogenic thickening within the sac. Fetal heartbeat is typically detectable by day 25 to 28, and this finding is the most secure early confirmation of a viable pregnancy. By day 30, the fetus has a recognizable shape with distinct head and trunk, and by day 35, limb buds and early organ differentiation become visible. The review by Lopate provides a detailed timeline of ultrasonographic fetal maturation that supports gestational age estimation when breeding dates are unknown [Lopate's ultrasonographic fetal maturation review](https://pubmed.ncbi.nlm.nih.gov/18534674/).

Operator skill is the dominant variable in early ultrasound diagnosis. A scan performed before day 20 may miss gestational sacs that are only 2 to 3 mm in diameter, and uterine fluid from other causes can mimic early pregnancy. Conversely, an experienced operator can often identify pregnancy at day 20 with high confidence. For the general practitioner, scheduling the first scan at day 25 to 28 reduces false negatives while still allowing early confirmation.

## Hormonal Assays: Relaxin and Progesterone

### Relaxin as a Pregnancy-Specific Marker

Relaxin assays are commercially available as point-of-care immunoassays and laboratory-based tests. The hormone becomes detectable in maternal serum from approximately day 22 to 25 after ovulation, and sensitivity approaches 100 percent when testing is performed after day 28. The test is specific for pregnancy because non-pregnant bitches do not produce placental relaxin. False positives are uncommon but have been reported in bitches with persistent luteal tissue or certain ovarian pathologies, so a positive result in a bitch with no other signs of pregnancy warrants ultrasound confirmation.

Timing is the principal limitation. A relaxin test performed before day 22 can yield a false negative because placental mass is insufficient to produce detectable hormone concentrations. The clinician should therefore record the ovulation date when known and delay testing until at least day 28 for maximum reliability. When ovulation timing is unknown, a negative relaxin result should be repeated 5 to 7 days later before pregnancy is excluded.

### Progesterone and the Limits of Luteal Assessment

Progesterone measurement has a defined but narrow role in pregnancy diagnosis. A single progesterone value confirms that ovulation has occurred if it exceeds the laboratory's threshold for luteal activity, typically 2 ng/mL. Serial measurements can confirm that the corpus luteum is functional, which is relevant when pregnancy loss is suspected. However, progesterone cannot confirm pregnancy because non-pregnant bitches in diestrus maintain luteal progesterone for 60 to 70 days, a duration that overlaps substantially with gestation.

The clinical value of progesterone in pregnancy diagnosis is therefore indirect. It establishes the timeline for scheduling ultrasound or relaxin testing, and it identifies bitches with luteal insufficiency that may require exogenous progestin support. The MSD Veterinary Manual provides species-specific guidance on interpreting progesterone concentrations in the context of canine reproductive staging [MSD Veterinary Manual professional edition](https://www.msdvetmanual.com/).

## Choosing a Diagnostic Strategy

The optimal approach depends on the clinical question. For simple confirmation of pregnancy, a single ultrasound at day 25 to 28 is efficient and provides additional information about fetal number and viability. For bitches with unknown breeding dates, ultrasound also enables gestational age estimation, which is critical for planning cesarean section or predicting whelping dates [Lopate's review of gestational age assessment](https://pubmed.ncbi.nlm.nih.gov/18534674/). For breeders who require early confirmation before day 25, a relaxin assay at day 22 with repeat testing at day 28 is a reasonable alternative, though it provides no fetal information.

When ultrasound and hormonal results conflict, the clinician should consider the timing of each test and the possibility of pregnancy loss. A positive relaxin result with no ultrasound findings may indicate an early pregnancy that was resorbed, or a scan performed too early. A negative relaxin result with ultrasound-confirmed pregnancy is rare after day 28 and should prompt assay validation or repeat testing. In all cases, the diagnostic plan should be documented and the owner counseled that no single test is infallible at every gestational stage.

## Ultrasonographic Findings by Gestational Stage

Ultrasound diagnosis of canine pregnancy rests on identifying discrete developmental landmarks in a predictable sequence. The earliest reliable finding is the embryonic vesicle, visible as an anechoic spherical structure within the uterine lumen. With a high-frequency transducer (7.5 to 10 MHz) and a skilled operator, vesicles can be detected as early as day 20 from the luteinizing hormone surge, though day 22 to 24 is a more practical target for routine confirmation. The Society for Theriogenology maintains professional resources on reproductive imaging standards and breeding management that can guide protocol design ([Society for Theriogenology resources](https://www.therio.org/)).

The zonary placental attachment becomes apparent between days 25 and 28. The embryo proper is visible within the vesicle by day 23 to 25, and fetal heartbeat is typically detectable by day 25 to 28. This heartbeat is the definitive confirmation of a viable pregnancy and distinguishes it from a nonviable vesicle or pseudopregnancy with uterine fluid. By day 28 to 30, the fetus has a discernible crown-rump length that can be measured reproducibly. Limb buds appear around day 30, and fetal movement is usually observed by day 32 to 35.

Skeletal ossification follows a craniocaudal gradient. The skull and spine mineralize first, visible on ultrasound as hyperechoic shadows by day 35 to 40. Ribs, scapulae, and long bones follow. Lopate's review of canine fetal maturation describes the use of both ultrasonographic and radiographic ossification timing to estimate gestational age when breeding dates are unknown ([Estimation of gestational age and assessment of canine fetal maturation using radiology and ultrasonography](https://pubmed.ncbi.nlm.nih.gov/18534674/)). This matters clinically because breeding dates alone are unreliable for due date prediction, particularly when mismating or silent estrus has occurred.

| Gestational landmark | Typical detection window | Clinical use |
|---|---|---|
| Embryonic vesicle | Day 20 to 24 | Earliest confirmation |
| Embryo proper | Day 23 to 25 | Confirms pregnancy |
| Fetal heartbeat | Day 25 to 28 | Confirms viability |
| Fetal movement | Day 32 to 35 | Viability assessment |
| Skull and spine ossification | Day 35 to 40 | Gestational age estimate |
| Ribs and long bones | Day 40 to 45 | Gestational age estimate |

## Fetal Count and Litter Size Estimation

Accurate fetal counting is one of the most clinically consequential tasks in canine pregnancy ultrasound. The bitch's bicornuate uterus and the overlapping of fetuses at later stages make counting error common. The optimal window for counting is day 28 to 35, when fetuses are small enough to be separated spatially but large enough to be identified reliably. After day 40, fetal overlap and uterine enlargement degrade counting accuracy substantially.

A systematic sweep is mandatory. Begin at the uterine body at the pelvic inlet and follow each horn cranially in a continuous transverse or sagittal plane. Count each fetus once by identifying its heartbeat or its distinct thorax. Record the count for each horn separately. A second sweep in the orthogonal plane reduces the risk of double counting. If the count differs between sweeps, repeat the examination instead of accepting the lower number.

The clinical limitation of ultrasound fetal counting must be stated plainly. Radiography after day 45, when the fetal skeleton is mineralized, provides a more reliable count in most cases. The MSD Veterinary Manual notes that radiographic counting is the preferred method for litter size estimation in late gestation ([MSD Veterinary Manual professional edition](https://www.msdvetmanual.com/)). Ultrasound remains superior for early confirmation and viability assessment, but a clinician who needs a precise count for whelping planning should combine early ultrasound with late radiography.

## Integrating Hormonal Assays into the Diagnostic Sequence

Relaxin and ultrasound are complementary instead of competing tests. Relaxin becomes detectable in maternal serum from day 22 to 25 after the luteinizing hormone surge, which coincides with the earliest ultrasound window. The two methods therefore have similar temporal onset, but they answer different questions. Relaxin confirms that placental tissue is present and secreting hormone. Ultrasound confirms that a fetus is present and viable.

The diagnostic sequence should be driven by the clinical question. When the question is simply "is this bitch pregnant?", a positive relaxin test at day 25 or later is sufficient in most cases. When the question is "is this pregnancy viable and how many fetuses are present?", ultrasound is required. When the question is "when will she whelp?", neither relaxin nor early ultrasound answers it directly. Gestational age estimation requires ultrasound measurement of fetal structures or late radiographic ossification assessment.

A negative relaxin test at day 25 should not be accepted as definitive. False negatives occur when testing is performed too early, when the assay is run incorrectly, or when placental mass is low. Repeat the test 3 to 5 days later before concluding the bitch is not pregnant. A positive relaxin test in a bitch with no detectable fetus on ultrasound raises the possibility of fetal death with retained placenta, or of an error in one of the two tests. Re-scan in 5 to 7 days to resolve the discrepancy.

## Equipment and Protocol Considerations

Transducer frequency is the single most important equipment decision. A 7.5 MHz linear or microconvex transducer is appropriate for most bitches under 25 kg. Larger breeds may require a 5 MHz transducer for adequate penetration, accepting reduced resolution. Obese bitches and those with significant abdominal fat degrade image quality at any frequency. Clip the ventral abdomen from the caudal mammary glands to the umbilicus. Acoustic coupling gel is mandatory, isopropyl alcohol alone is inadequate.

Patient positioning affects diagnostic yield. Dorsal recumbency with the hindlimbs pulled caudally is standard. Some deep-chested or anxious bitches tolerate lateral recumbency better, and the gravid uterus falls toward the dependent side, which can improve access. The urinary bladder should be assessed first as a landmark. The uterine body lies dorsal to the bladder neck, and the horns extend cranially from there.

Documentation should include the date relative to the luteinizing hormone surge or breeding, the number of vesicles or fetuses per horn, the presence and rate of fetal heartbeats, and any measured fetal structures used for gestational age estimation. Store representative images or video loops in the medical record. This documentation supports later comparison if the pregnancy is re-scanned, and it provides a baseline if fetal loss is suspected.

## Decision Points and When the Strategy Changes

The correct diagnostic strategy changes with the clinical scenario. A breeding management program with timed ovulation uses ultrasound at day 25 to 28 to confirm pregnancy and day 28 to 35 to count fetuses. A rescue or mismating scenario requires earlier diagnosis, and relaxin testing at day 25 is the most practical first step. A bitch with suspected fetal death needs ultrasound, not relaxin, because relaxin can remain positive for days after fetal loss.

Singleton pregnancies present a specific trap. A single fetus is easy to miss on a cursory scan, and the bitch may be incorrectly diagnosed as nonpregnant. The converse error also occurs. A bitch with pyometra or cystic endometrial hyperplasia can have uterine fluid that mimics embryonic vesicles. The presence of a fetal heartbeat is the only reliable discriminator. If no heartbeat is seen, do not diagnose pregnancy.

The evidence base for some claims in canine pregnancy diagnosis is limited. Much of the published timing data derives from small studies or clinical series, and breed size affects both the visibility of early structures and the accuracy of gestational age estimates. The review by Lopate acknowledges these limitations and recommends that clinicians establish their own reference ranges where possible ([Estimation of gestational age and assessment of canine fetal maturation using radiology and ultrasonography](https://pubmed.ncbi.nlm.nih.gov/18534674/)). A clinician who records gestational measurements from confirmed pregnancies and correlates them with actual whelping dates will build a more reliable local reference than any published table.

## Recognized Complications and Early Detection

Pregnancy diagnosis in the bitch carries specific failure modes that the clinician must anticipate. The most consequential is the false-negative diagnosis performed too early. Ultrasonography before day 20 of gestation can miss a pregnancy because the embryonic vesicles are small and easily overlooked, particularly with inexperience or suboptimal transducer placement. A negative scan before day 25 should therefore be repeated within 5 to 7 days before the owner is counselled that the bitch is not pregnant.

Embryonic resorption is a recognized complication that can convert an initially positive diagnosis into a negative outcome. Early pregnancy loss may be detected ultrasonographically as loss of fetal heartbeat, collapse of the gestational sac, or progressive reduction in chorionic cavity size. Serial scanning is the only reliable method to distinguish resorption from a primary non-pregnancy. When a heartbeat has been documented and then disappears on a subsequent examination, the diagnosis of embryonic death is secure.

Pregnancy loss after day 45 is less common but more clinically significant because it may require intervention. Fetal death at this stage is recognized by absence of cardiac motion, loss of fetal tone, and eventually by hyperechoic gas within the uterine lumen as autolysis progresses. The clinician should also recognize that a singleton pregnancy may be missed on palpation but is readily identified by ultrasound, and that an apparently non-pregnant bitch with a persistent luteal phase may have undergone complete resorption instead of never having conceived.

## Common Errors and Corrective Action

Less experienced clinicians make characteriztic errors that are largely avoidable with disciplined technique. The most frequent is mistaking the urinary bladder for a gestational sac. The bladder is anechoic, thin-walled, and located caudally, whereas gestational sacs are spherical, thick-walled, and distributed along the uterine horns. The discriminating check is to sweep the transducer cranially from the bladder, gestational sacs appear as multiple discrete structures, not a single large cavity.

A second common error is misidentifying the uterine body as a gestational sac during early pregnancy. The uterine body appears as a tubular, fluid-filled structure in the caudal abdomen and can be confused with a single vesicle. The corrective action is to scan both uterine horns in their entirety and to confirm that the suspected sac has a distinct trophoblastic wall with a lumen that is separate from the uterine lumen.

A third error is the overinterpretation of a single progesterone measurement. Progesterone confirms luteal activity, not pregnancy, and a bitch with a prolonged luteal phase from cystic endometrial hyperplasia or pyometra can have elevated progesterone without being pregnant. The discriminating check is to combine progesterone with relaxin measurement or ultrasound, never to rely on progesterone alone.

| Observation | Likely Cause | Discriminating Check |
| --- | --- | --- |
| Single large anechoic structure caudally | Urinary bladder | Sweep cranially, bladder is solitary, gestational sacs are multiple |
| Tubular fluid-filled structure in caudal abdomen | Uterine body | Trace both horns, confirm distinct trophoblastic wall |
| Positive progesterone, negative ultrasound | Non-pregnant luteal phase or early resorption | Repeat ultrasound in 5 to 7 days, measure relaxin |
| Fetal heartbeat absent after day 25 | Embryonic death | Confirm with repeat scan 24 to 48 hours later |
| Decreasing chorionic cavity size | Resorption | Serial scanning, document loss of fetal heartbeat |

## Limitations of the Current Evidence

The evidence base for canine pregnancy diagnosis is narrower than for other domestic species. Much of the published guidance on gestational age estimation derives from a limited number of studies, and the review by Lopate on gestational age assessment using radiology and ultrasonography notes that breeding dates alone do not provide due dates with adequate accuracy. This remains a practical limitation because many bitches are presented without precise ovulation timing.

Expert opinion still differs on the optimal timing for the first confirmatory ultrasound. Some authorities recommend scanning at day 21 to 25, while others advocate waiting until day 25 to 28 to reduce the false-negative rate. The Society for Theriogenology provides professional resources on reproductive management, but specific recommendations vary by institution and clinical context. The MSD Veterinary Manual similarly offers general guidance without resolving the timing debate.

The accuracy of fetal counts is another area where the evidence is contested. Ultrasound underestimates litter size in large litters, and the clinician should communicate this limitation to the owner instead of promising a precise number. Radiography after day 45 remains the most accurate method for fetal counting, but it does not confirm viability.

## Referral, Consultation, and Reporting

Most pregnancy diagnoses are completed in general practice without specialist input. Referral to a theriogenologist or veterinary radiologist is warranted when the ultrasound examination is technically difficult, when fetal abnormalities are suspected, or when the clinician cannot confidently determine gestational age for planned cesarean section. A bitch with a suspected singleton pregnancy and no breeding dates is a candidate for specialist assessment because accurate gestational age is critical for surgical planning.

Laboratory involvement is indicated when relaxin assay results are discordant with ultrasound findings. A positive relaxin with a negative ultrasound may indicate very early pregnancy, while a negative relaxin with a positive ultrasound suggests assay error or a rare false-negative result. The laboratory should be consulted to confirm the assay was performed correctly and to discuss the expected performance characteriztics of the specific test.

Regulatory reporting is rarely required for routine pregnancy diagnosis. However, the clinician should be aware that certain jurisdictions have reporting requirements for procedures performed on animals used in research or for endangered species. The WOAH terrestrial animal health standards address animal health surveillance and welfare, and the clinician working with non-domestic canids should consult these standards. The AVMA practice resources provide additional guidance on professional obligations in unusual cases.

## Frequently Asked Questions

### How Reliable Is a Negative Relaxin Test Before Day 30?

A negative relaxin result before day 30 does not exclude pregnancy. Relaxin is produced by the placenta, and circulating concentrations may not reach assay detection thresholds until implantation is firmly established. Most commercial point-of-care tests are validated for use from day 28 or 30 after breeding, and testing earlier risks a false negative. If clinical suspicion remains high, repeat the assay after day 30 or proceed directly to ultrasonography. Ultrasound can identify a gestational sac as early as day 20 and an embryo by day 23, making it the preferred method when earlier confirmation is required. The [Society for Theriogenology resources](https://www.therio.org/) provide guidance on assay timing and interpretation.

### Can Ultrasound Confirm Non-Pregnancy, or Only Pregnancy?

Ultrasound confirms pregnancy when embryonic vesicles or fetuses are visualized, but it cannot definitively confirm non-pregnancy at a single early examination. A scan performed before day 20 may miss small vesicles, and uterine fluid can be mistaken for early pregnancy or, conversely, early pregnancy can be mistaken for cystic endometrial hyperplasia. A negative scan before day 25 should be repeated at day 30 or later. At that stage, the absence of gestational sacs with a normal uterine appearance supports non-pregnancy, particularly when combined with a negative relaxin assay. The [MSD Veterinary Manual](https://www.msdvetmanual.com/) notes that diagnostic confidence increases when imaging is timed to known gestational stages.

### What Is the Minimum Ultrasound Equipment Needed for Reliable Diagnosis?

A high-frequency linear or microconvex transducer operating at 7.5 to 10 MHz is sufficient for early canine pregnancy diagnosis. Lower-frequency transducers, such as those used for large animal abdominal work, may fail to resolve the small embryonic vesicles before day 25. The bitch is scanned in dorsal recumbency with the transducer placed caudal to the umbilicus, and the urinary bladder serves as an acoustic window. If only a lower-frequency probe is available, delay the scan until day 28 or later, when the vesicles are larger and more readily identified. Color Doppler is not required for diagnosis, though it can aid in confirming embryonic heartbeat when cardiac motion is ambiguous on B-mode.

### How Should Results Be Recorded for Serial Comparison?

Record the date relative to the first breeding and to ovulation if known, the transducer frequency, and the specific findings at each scan: number of gestational sacs, crown-rump length, presence of a heartbeat, and any placental or fetal abnormality. Use a standardized worksheet so that serial examinations are directly comparable. Include the relaxin assay result with the assay brand and the day of gestation on which it was performed. This record supports gestational age estimation and informs decisions about timing of radiographs for fetal count later in gestation. The [Society for Theriogenology resources](https://www.therio.org/) offer templates for reproductive records that can be adapted to practice use.

### How Does the Diagnostic Approach Differ in a Practice Without Ultrasound?

When ultrasound is unavailable, relaxin assay combined with careful abdominal palpation is the practical alternative. Palpation is reliable between days 28 and 35, when the uterine swellings are distinct, but becomes unreliable after day 40 as the uterus enlarges and the fetuses become less individually distinguishable. Relaxin testing from day 30 onward provides a pregnancy-specific result independent of examiner skill. Radiography can confirm pregnancy from day 45 when fetal skeletons mineralise, and it also permits fetal counting. The [AVMA practice resources](https://www.avma.org/resources-tools) note that referral for ultrasound should be considered when early confirmation is needed for breeding management decisions.

### How Should Discordant Ultrasound and Relaxin Results Be Interpreted?

Discordance arises most often from timing errors. A positive relaxin with a negative ultrasound before day 25 usually reflects scanning too early, and the scan should be repeated. A negative relaxin with a positive ultrasound after day 30 is more concerning and may indicate assay failure, sample handling error, or a rare placental abnormality. Repeat both tests within 48 to 72 hours. If the ultrasound finding is unequivocal, such as a visible embryo with a heartbeat, the imaging result takes precedence. The [MSD Veterinary Manual](https://www.msdvetmanual.com/) advises that no single test is infallible and that repeat testing resolves most discrepancies. Document the discordance and the repeat results in the medical record.

## Related Clinical & Scientific Guides

* [Diagnostic Approach to Canine Infertility in the Bitch](/knowledge/veterinary-medicine/theriogenology/diagnostic-approach-to-canine-infertility-in-the-bitch)
* [Canine Neonatal Resuscitation: Protocol and Monitoring](/knowledge/veterinary-medicine/theriogenology/canine-neonatal-resuscitation-protocol-monitoring)
* [Equine Breeding Soundness Examination of the Stallion](/knowledge/veterinary-medicine/theriogenology/equine-breeding-soundness-examination-of-the-stallion)


## References and Further Reading

- [Estimation of gestational age and assessment of canine fetal maturation using radiology and ultrasonography: a review.](https://pubmed.ncbi.nlm.nih.gov/18534674/). 2008.
- [Ovarian cancer (review). Etiology, diagnosis, prognosis, surgery, radiotherapy, chemotherapy and endocrine therapy.](https://pubmed.ncbi.nlm.nih.gov/3291746/). 1988.
- [Rhombencephalosynapsis and related anomalies: a neuropathological study of 40 fetal cases.](https://pubmed.ncbi.nlm.nih.gov/19057916/). 2009.
- [Alcohol as a teratogen: a decade of research in review.](https://pubmed.ncbi.nlm.nih.gov/3322304/). 1987.
- [Endangered wolves cloned from adult somatic cells.](https://pubmed.ncbi.nlm.nih.gov/17386020/). 2007.
- [In vivo effect of interleukin-1beta and interleukin-1RA on oocyte cytoplasmic maturation, ovulation, and early embryonic development in the mare.](https://pubmed.ncbi.nlm.nih.gov/15972098/). 2005.
- [Society for Theriogenology Resources](https://www.therio.org/). Society for Theriogenology.
- [MSD Veterinary Manual, Professional Edition](https://www.msdvetmanual.com/). MSD Veterinary Manual.
- [American Veterinary Medical Association Practice Resources](https://www.avma.org/resources-tools). American Veterinary Medical Association.

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- [Feline Pregnancy Diagnosis: Palpation, Ultrasound, and Hormonal Assays](/knowledge/veterinary-medicine/theriogenology/feline-pregnancy-diagnosis-palpation-ultrasound-hormonal-assays)
- [Ovine Pregnancy Diagnosis: Transabdominal Ultrasound and Management](/knowledge/veterinary-medicine/theriogenology/ovine-pregnancy-diagnosis-transabdominal-ultrasound-management)
- [Pregnancy Diagnosis in Cattle: Methods and Accuracy](/knowledge/veterinary-medicine/theriogenology/pregnancy-diagnosis-in-cattle-methods-and-accuracy)
- [Differential Diagnosis of Canine Vaginal Discharge](/knowledge/veterinary-medicine/theriogenology/differential-diagnosis-of-canine-vaginal-discharge)
- [Equine Pregnancy Diagnosis: Methods and Clinical Application](/knowledge/veterinary-medicine/theriogenology/equine-pregnancy-diagnosis-methods-and-clinical-application)

> This article is educational professional reference material for veterinary audiences. It is not a substitute for veterinary diagnosis, individual clinical judgment, current product labeling, or applicable regulatory requirements.