Preparing for Blood Test Athletes: Fasting, Timing, and Training Considerations

Introduction

Many of my athletic patients follow highly structured training programs. Their routines often include near-daily workouts, carefully planned nutrition, regular supplement use, and strict schedules. However, I frequently see how this structure can conflict with optimal conditions for blood testing. A hard training session the day before, a poorly timed blood draw, or a biotin supplement that wasn’t mentioned can each transform an accurate baseline into a clinically misleading snapshot.

In practice, I’ve also noticed that most laboratory instructions provided by clinics are very limited. Typically, they mention only basic requirements — such as fasting or taking certain tests in the morning. However, they rarely address factors that are highly relevant for athletes, such as recent training load or, in female athletes, the phase of the menstrual cycle.

In reality, there is very little practical guidance specifically designed for athletes, even though these factors can clearly influence test results. This gap is one of the main reasons I wrote this guide — to provide clear, practical guidance on preparing for blood test athletes can actually follow in real-world conditions.

In this guide, I walk through what the evidence actually says about preparing for athletic blood work — including fasting requirements, time-of-day standardization, training timing, hydration, supplement interference, and menstrual cycle considerations for female athletes. My goal is not just to help you get a single accurate result, but to create reproducible testing conditions you can follow every time, so your data tells a reliable story over months and years.

Preparing for Blood Test Athletes: Why It Matters

One important point I often explain to my patients is how reference ranges are actually defined. Most laboratory reference values are constructed so that 95% of a general population falls within that range — and that population is predominantly sedentary or only moderately active.

This creates a challenge for athletes. They often operate outside these norms for markers such as hematocrit, creatine kinase (CK), and resting cortisol. When a biomarker is influenced by training, an athlete can fall outside the reference range despite being physiologically normal. In other words, what appears “abnormal” on paper may simply reflect adaptation to training rather than pathology.

Another way I explain this is that much of the preparation before blood testing is, in practice, an attempt to temporarily minimize the acute effects of training — so that results are more comparable to standard reference populations. We are not trying to “normalize” the athlete, but rather to reduce short-term physiological fluctuations caused by recent exercise, nutrition, or timing. Without this step, it becomes difficult to distinguish between true baseline physiology and transient training effects. Even with careful preparation, however, athletes do not fully fit into standard reference frameworks, and some deviations may still reflect normal adaptation rather than disease.

Research published in Sports Medicine on blood biomarker monitoring in elite athletes is explicit: preanalytical conditions are a major source of measurement error, and evidence-based recommendations include avoiding vigorous exercise the day before collection, fasting overnight, and standardizing the time of day to account for diurnal hormonal variation [1]. The same authors note that these conditions are particularly difficult to satisfy in elite sport settings — making a systematic personal protocol all the more important for athletes who test themselves.

Another challenge I see in practice is that there is no true standardization specifically designed for athletes. Creating one is inherently difficult, because different sports — and even different training styles within the same sport — affect physiology in very different ways.

Because of this, I often have to interpret laboratory results in athletes on an individual basis. I can’t rely solely on standard reference ranges. Instead, I need to consider the athlete’s training load, sport, and current phase of training, and interpret results in that context.

In many cases, this also means that athletes benefit from working with a clinician who understands these sport-specific physiological adaptations — rather than relying purely on standard laboratory interpretations.

It is also worth noting that some markers carry population-level variation. Benign ethnic neutropenia — a lower baseline neutrophil count seen in some individuals of African, Middle Eastern, or West Indian descent — is a normal genetic variant and not a sign of immune suppression [16]; an athlete in this group should not be flagged for low immunity based on neutrophil counts that fall within their healthy genetic baseline.

Fasting Requirements: What Actually Needs a Fast, and What Doesn’t

Not every test on an athletic blood panel requires fasting — but several key markers do, and confusing the two is one of the most common preparation errors. In clinical settings, fasting requirements are usually clearly indicated — often with an “f” prefix — and clinicians typically provide proper instructions. However, when athletes order commercial lab panels themselves, guidance may be limited or unclear, and fasting is sometimes missed altogether.

Tests that genuinely require fasting (10-12 hours, water permitted):

Fasting glucose and the oral glucose tolerance test (OGTT) require fasting because food intake directly elevates blood sugar, creating results that reflect a postprandial state rather than your baseline metabolic function. Triglycerides within a lipid panel are the most food-sensitive component: elevated levels persist for hours after a fat-containing meal [2]. A joint consensus statement from the European Atherosclerosis Society and the European Federation of Clinical Chemistry and Laboratory Medicine confirmed that while nonfasting lipids are now acceptable for general cardiovascular risk screening, fasting remains preferable for accurate triglyceride quantification [2]. If you eat a high-fat meal the night before, expect artificially elevated triglyceride readings.

Iron studies — including serum iron, TIBC, and transferrin saturation — are also conventionally drawn fasting, though the evidence on exactly how much fasting matters is more nuanced than commonly understood (see the section on timing below).

In practice, iron status is often assessed using basic tests such as a complete blood count and ferritin, which do not require fasting. However, in more comprehensive panels, additional markers such as TIBC and serum iron are frequently included — and these are typically measured under fasting conditions.

Tests that do not require fasting:

Hemoglobin A1c (HbA1c) measures a three-month average and is completely unaffected by recent food intake. Ferritin, a marker athletes use to assess iron stores, is not substantially altered by short-term fasting status in otherwise healthy individuals, though inflammation from recent hard training can artificially elevate it regardless of fasting (discussed in the training section). Complete blood count (CBC) markers — hemoglobin, hematocrit, RBC, WBC — are not significantly affected by fasting. Thyroid function tests (TSH, fT4) do not require fasting, though they are sensitive to supplement interference, discussed below.

In practice, it is often possible to assess a patient’s “fatigue panel” without fasting, which means testing can frequently be done already at the first visit. These panels typically include a complete blood count, thyroid function tests, ferritin, vitamin D, and similar baseline markers.

Time of Day: Why Hormonal Tests Especially Demand Morning Draws

The time of blood collection is arguably more important for hormonal markers than fasting status is. Several key markers athletes track follow pronounced circadian rhythms, and drawing at the wrong time of day can produce clinically misleading values.

Testosterone has the strongest time-of-day dependence of any commonly tested marker. Research from the Boston Area Community Health Survey, published in the Journal of Clinical Endocrinology and Metabolism, studied 66 men aged 30–80 across morning and afternoon draws. In men aged 30–40, testosterone was 20–25% lower at 16:00 than at 08:00 [3]. The clinical impact was significant: half of the men with at least one afternoon testosterone measurement below the hypogonadal threshold of 300 ng/dL had entirely normal morning values across three separate draws [3].

This has clear clinical implications. In recent years, there has been a growing trend of evaluating fatigue in men with testosterone testing, sometimes leading to a diagnosis of hypogonadism based on a single measurement taken under suboptimal conditions. In such cases, an afternoon sample may falsely suggest low testosterone, increasing the risk of overdiagnosis and unnecessary treatment. This is particularly relevant because testosterone replacement therapy, when used without clear indication, can suppress endogenous production and impair fertility.

Cortisol follows a well-characterized diurnal rhythm driven by the hypothalamic-pituitary-adrenal (HPA) axis: circulating concentrations peak at or shortly after the habitual sleep-wake transition and decline progressively to a nadir during the late evening and early night [4]. This is why cortisol — and the testosterone-to-cortisol ratio used in overtraining assessment — must be drawn at a standardized morning time to be interpretable. Afternoon cortisol values are physiologically lower and cannot be meaningfully compared to morning reference ranges.

In routine clinical practice, however, cortisol is not commonly measured outside of specific endocrine evaluations. In most healthcare settings, including Finland, testing is typically performed under clearly defined protocols and requires a clinician’s referral. It is therefore less frequently included in standard or commercial laboratory panels without a specific clinical indication.

Serum iron shows more variable diurnal behavior than is often assumed. The Mayo Clinic study by Dale et al. (2002), which measured serum iron, TIBC, transferrin saturation, and ferritin in 20 volunteers at 08:00, 12:00, and 16:00, found that while statistically significant differences in mean values existed across time points, no consistent individual diurnal pattern was observed — morning levels were higher than afternoon levels in only half of subjects [5]. The authors concluded that restricting iron draws to morning hours does not reliably improve test reliability. A large population-based analysis of over 276,000 samples found iron levels were relatively elevated and stable from approximately 08:00 to 15:00, peaking around 11:00 to 12:00 in adults [6]. The practical implication: morning is still preferable for iron panel standardization, particularly for serial testing, but the rigid 08:00 cut-off is less critical than once believed.

I also see that some other tests come with timing considerations, although they are less strict. For example, in our lab, thyroid function tests are recommended to be taken before 16:00 to ensure consistency and comparability of results. In practice, I can usually get these tests done without issues, unless the patient comes in for an evening appointment

Training Timing: The 48-Hour Rule

Acute exercise causes transient elevations in multiple blood markers, several of which can remain outside normal ranges for 24–72 hours post-exercise. Testing too close to hard training is one of the most common sources of misinterpretation in athletic populations.

Creatine kinase (CK) is the most exercise-sensitive commonly tested marker. A case report documented a 22-year-old athlete presenting with CK of 3,600 U/L following intense combined weightlifting and endurance training the previous day — high enough to trigger investigation for cardiac or muscular pathology — which normalized completely within two days of rest [7]. For trained athletes, CK may return toward baseline within 24 hours of moderate exercise, but following heavy eccentric loading or competition it can remain elevated for several days. If you’ve recently run a marathon, see our detailed guide on post-marathon blood work normalization for expected recovery timelines across specific markers.

With CK, however, I always take a more careful approach. I make sure to ask about symptoms such as unusual muscle pain or dark-colored urine. While exercise can significantly elevate CK levels, rhabdomyolysis must always be kept in mind and ruled out when clinically suspected.

ALT and AST, traditionally considered liver enzymes, are also found in skeletal muscle and can be transiently elevated post-exercise through the same mechanism of muscle microtrauma — which has led to inappropriate liver disease workup in athletes whose training history wasn’t taken into account [7]. When I see more significant or persistent increases, they are more often related to other causes — such as alcohol use or fatty liver disease — which, while generally less common in athletic populations, still need to be considered.

Leukocyte count increases significantly during and immediately after intense exercise, likely driven by stress-related and catecholamine-mediated immune cell mobilization, and may exceed normal reference limits in the post-exercise period[8]. An early study of 19 serum and hematological parameters in runners before and after a 13-mile race found post-exertional values above the upper limit of normal in 71% of subjects for leukocyte count and 93% for CK[8].

Transient leukocytosis, on the other hand, rarely leads to immediate interventions in my practice. In most cases, I simply recheck the value. While exercise can elevate leukocyte counts, there are many other common causes as well — including infection, physiological stress, and medications such as corticosteroids.

Ferritin can be falsely elevated following hard training due to its acute phase reactant properties — exercise-induced inflammation raises ferritin independent of actual iron stores [9]. Similarly, C-reactive protein (CRP), which athletes sometimes track alongside iron panels to contextualize ferritin readings, is elevated following hard training for at least 24 hours [9].

Elevated CRP, on the other hand, usually warrants follow-up. In my practice, I typically recheck it, and if the level is clearly elevated — for example above 30 mg/L — I start to look more closely for an underlying cause, such as an infection.

Ideally, I recommend scheduling blood work at least 48 hours after any high-intensity session, long endurance bout, or competition. In practice, this can be challenging for many athletes, as they rarely go two days without training.

Hydration Status: The Often-Overlooked Variable

Dehydration and exercise-induced plasma volume shifts concentrate blood markers through hemoconcentration — the reduction in plasma volume increases the relative concentration of cellular and protein components, which can transiently elevate concentration-dependent markers such as hemoglobin and hematocrit. Research on elite kayak-canoe athletes found hemoglobin concentration reached a median of 17.4 g/dL and hematocrit 53.5% at peak exercise loading — well above resting baselines — before returning to normal levels within 30 minutes of recovery [10]. For athletes whose resting hydration is also compromised (weight-cutting protocols, heat training camps, travel), the same concentrating effect can persist at rest.

For athletes whose testing includes hematocrit, hemoglobin, or any concentration-dependent markers, arriving at the draw in a euhydrated state is essential. Pre-analytic hydration protocols referenced in elite athlete blood monitoring guidelines [1] emphasize maintaining a euhydrated state, with normal fluid intake in the hours leading up to the blood draw. This is also relevant for athletes concerned aboutdilutional pseudoanemia (sports anemia) — a condition where increased plasma volume from training makes hemoglobin appear artificially low.

I also sometimes see that fasting morning samples can appear hemoconcentrated, particularly in individuals who have not only fasted but also avoided fluids for an extended period. Although fasting technically allows water intake, many people drink very little — and since food normally contributes a meaningful amount of daily fluid intake, this can lead to relative dehydration. In these cases, markers such as hemoglobin and hematocrit may appear artificially elevated due to reduced plasma volume.

Supplements to Avoid Before Testing

Biotin (vitamin B7) is found in many sports-focused multivitamins and is widely marketed for hair, skin, and nail health at doses ranging from 5–20 mg — far exceeding the recommended dietary intake of 30 µg/day. At these supplemental doses, biotin can cause significant interference with a broad range of immunoassays that use biotin-streptavidin binding in their design.

In this context, I think it is important to emphasize that biotin interference is primarily a laboratory assay artifact rather than a true reflection of altered physiology. In these cases, the patient’s actual hormone or biomarker levels may be unchanged, but the immunoassay reports a falsely low or falsely high result due to biotin–streptavidin interference.

The clinical impact is well-documented. A prospective study published in Thyroid (Ylli et al. 2021) administered 10 mg/day of biotin to 13 adult subjects and found significant interference with TSH, fT4, and TT3 measurements on multiple laboratory platforms, with maximal interference occurring two hours after ingestion [11]. The FDA has issued safety communications specifically about biotin interference with troponin assays, noting at least one potentially preventable death linked to a false-negative troponin result in the context of high biotin intake [12].

For athletes, the markers most likely to be affected by biotin supplementation include TSH (falsely low on sandwich assays), free T4 and free T3 (falsely elevated on competitive assays) — see our full guide on thyroid function in athletes — as well troponin[12].

Biotin washout time is dose- and assay-dependent. Even moderate supplement doses (e.g., 5–10 mg/day) can interfere with certain assays, and the duration of interference may vary depending on the laboratory platform[11][12].

I also occasionally see patients who use creatine supplements, which can lead to elevated creatinine levels. These individuals often have higher muscle mass as well, which further contributes to the finding. Importantly, this does not necessarily reflect impaired kidney function, but rather increased creatinine production from exogenous creatine and muscle metabolism. In such cases, I usually advise pausing creatine supplementation and repeating the test later to clarify the result.

Menstrual Cycle Timing for Female Athletes

For female athletes tracking iron status, menstrual cycle phase at the time of blood draw introduces meaningful variability in several markers, and failing to account for it can lead to misclassification of iron status. This is particularly relevant for the conditions discussed in our guide on RED-S and the Female Athlete Triad, where accurate iron assessment is central to diagnosis.

Importantly, these variations do not reflect iron status alone. They represent a combination of true changes in iron balance — such as menstrual blood loss — and hormonally driven physiological fluctuations affecting iron distribution, plasma volume, and regulatory pathways. As a result, a single measurement taken at a different phase of the menstrual cycle may not accurately reflect underlying iron stores, but rather the timing of the test.

A large NHANES-based analysis of 1,712 women found hemoglobin, transferrin saturation, and serum ferritin were lowest during the menstrual phase and highest during the luteal and late luteal phases (Hb 130 vs. 133 g/L; transferrin saturation 21.2% vs. 24.8%; ferritin 17.2 vs. 24.0 µg/L, all p<0.05)[13]. A recent NHANES analysis of 1,484 women across all cycle phases confirmed serum iron peaked in the early/mid-luteal phase and was lowest during menstruation, whilesoluble transferrin receptor was highest during menstruation, reflecting increased iron requirements[14].

Research specifically in endurance-trained athletes reinforces this. Alfaro-Magallanes et al. (2022) found iron and transferrin saturation in the early follicular phase were substantially lower than in the late follicular and mid-luteal phases in 21 eumenorrheic athletes, with the authors explicitly noting that testing during the early follicular phase may mislead the determination of iron status [15]. Notably, ferritin appears more stable across the cycle than serum iron or TSAT [14][15], making it a more cycle-robust marker for iron stores, though not immune to inflammation-driven fluctuation. For a full picture of how to interpret the complete iron panel in this context, see Iron Panel Interpretation for Female Athletes.

For consistent serial iron status monitoring in female athletes, testing should ideally be standardized to the same phase of the menstrual cycle — most practically the mid-to-late follicular phase (days 7–14 of a standard 28-day cycle), while avoiding the first 5 days of menstruation when iron markers are most likely to be suppressed. Cycle day should always be recorded alongside results.

In practice, I often see this aspect being overlooked. Iron status is frequently assessed without considering menstrual cycle timing, which can lead to misinterpretation of results. In my view, greater awareness of this variability would improve clinical decision-making. At a minimum, I aim to repeat follow-up measurements in the same cycle phase as previous tests to allow for meaningful comparison.

I also find that this becomes more challenging in athletes using hormonal contraception, particularly in those without a regular menstrual cycle, where phase-based timing is no longer applicable.

Conclusion

Accurate blood work in athletes begins long before the sample is taken. In practice, results are shaped not only by the test itself, but also by what happens in the hours and days beforehand — training load, nutrition, hydration, supplement use, time of day, and, in female athletes, the phase of the menstrual cycle. If these factors are not accounted for, results may appear precise but fail to reflect true baseline physiology.

This is why preparation is especially important in athletic populations. The goal is not to force athletes into standard reference ranges, but to reduce avoidable short-term variability so that results become more interpretable and comparable over time. Even with optimal preparation, athletes often require more individualized interpretation, as training itself produces meaningful physiological adaptations.

When testing conditions are standardized, blood work becomes far more than a one-time snapshot. It becomes a practical tool for monitoring health, guiding recovery, and supporting long-term performance.

References

1. https://pmc.ncbi.nlm.nih.gov/articles/PMC10197055/
2. https://pmc.ncbi.nlm.nih.gov/articles/PMC4929379/
3. https://pmc.ncbi.nlm.nih.gov/articles/PMC2681273/
4. https://pmc.ncbi.nlm.nih.gov/articles/PMC8813037/
5. https://pubmed.ncbi.nlm.nih.gov/12090432/
6. https://www.sciencedirect.com/science/article/abs/pii/S0009912017304526
7. https://pmc.ncbi.nlm.nih.gov/articles/PMC11498664/
8. https://pubmed.ncbi.nlm.nih.gov/7072633/
9. https://pmc.ncbi.nlm.nih.gov/articles/PMC5640004/
10. https://pmc.ncbi.nlm.nih.gov/articles/PMC9803156/
11. https://pubmed.ncbi.nlm.nih.gov/34042535/
12. https://pmc.ncbi.nlm.nih.gov/articles/PMC6802814/
13. https://pubmed.ncbi.nlm.nih.gov/8237879/
14. https://pmc.ncbi.nlm.nih.gov/articles/PMC12681887/
15. https://pubmed.ncbi.nlm.nih.gov/36129579/
16. https://pmc.ncbi.nlm.nih.gov/articles/PMC6702066/