Boronate affinity HbA1c principle is the chemical basis behind one of the most widely used methods for measuring glycated haemoglobin in diabetes care. This approach exploits the selective interaction between phenylboronic acid and cis-diol groups on glucose molecules bound to haemoglobin, enabling accurate separation and quantification of glycated haemoglobin fractions. Understanding how this chemistry works — and how it compares to other analytical methods such as ion-exchange HPLC — is essential for clinicians, laboratory staff, and anyone involved in diabetes diagnosis or monitoring within the NHS. This article explains the science, clinical application, and quality considerations surrounding boronate affinity HbA1c testing.

What Is HbA1c and Why It Matters in Diabetes Monitoring

HbA1c reflects average blood glucose over the preceding two to three months due to non-enzymatic glycation of haemoglobin throughout the red cell lifespan. An HbA1c of 48 mmol/mol or above is diagnostic of type 2 diabetes per NICE NG28.

Haemoglobin A1c (HbA1c) is a form of haemoglobin that forms when glucose in the bloodstream binds irreversibly to the beta-chain of haemoglobin A. This process, known as non-enzymatic glycation, occurs continuously throughout the lifespan of a red blood cell — approximately 120 days. Because the degree of glycation reflects average blood glucose concentrations over the preceding two to three months, HbA1c provides a reliable, long-term indicator of glycaemic control that a single fasting glucose measurement cannot offer.

In clinical practice, HbA1c is central to both the diagnosis and ongoing management of diabetes mellitus. According to NICE guidelines (NG28), an HbA1c of 48 mmol/mol (6.5%) or above is diagnostic of type 2 diabetes. In asymptomatic adults, this should be confirmed with a repeat HbA1c measurement; in adults with typical symptoms of diabetes, a single result at or above this threshold may be sufficient. Levels between 42–47 mmol/mol indicate non-diabetic hyperglycaemia (also described as a high risk of developing type 2 diabetes), and these individuals should be offered lifestyle advice and regular review. The term 'prediabetes' is not preferred in UK clinical practice; 'non-diabetic hyperglycaemia' is the recommended terminology.

Importantly, HbA1c is not appropriate for diagnosis in several circumstances, including:

• Children and young people

• Pregnancy

• Suspected type 1 diabetes or rapidly evolving diabetes of any type

• Conditions that alter red cell turnover, such as haemolytic anaemia, iron deficiency anaemia, or within two months of a blood transfusion

In these situations, plasma glucose measurement (fasting or random, with or without an oral glucose tolerance test) should be used instead, in line with NICE NG28 and WHO 2011 guidance.

For people already living with diabetes, regular HbA1c monitoring — typically every three to six months — helps clinicians assess whether treatment targets are being met and guides decisions about medication adjustment. Large-scale trials such as the UKPDS demonstrated that sustained reductions in HbA1c are associated with meaningful decreases in the risk of microvascular complications, including diabetic retinopathy, nephropathy, and neuropathy. This evidence base underpins the NHS's continued investment in standardised, accurate HbA1c measurement across primary and secondary care settings.

The Boronate Affinity Principle Explained

Phenylboronic acid forms a reversible covalent ester bond with cis-diol groups on glycated haemoglobin at alkaline pH, selectively retaining glycated species on a boronate resin whilst non-glycated haemoglobin passes through unbound.

Boronate affinity chromatography exploits a well-characterised chemical interaction between phenylboronic acid and the cis-diol groups found on glucose molecules that have become covalently attached to haemoglobin. When glucose glycates haemoglobin, it forms a stable ketoamine linkage — specifically at the N-terminal valine of the haemoglobin beta-chain. The resulting glycated haemoglobin retains adjacent hydroxyl (–OH) groups in a cis configuration on the attached sugar moiety, which are highly reactive with boronic acid derivatives.

At a slightly alkaline pH, phenylboronic acid forms a reversible, covalent ester bond with these cis-diol groups. This selective binding is the cornerstone of the boronate affinity principle: glycated haemoglobin binds to the boronate-functionalised resin, whilst non-glycated haemoglobin passes through unretained. The bound glycated fraction can then be eluted by lowering the pH or introducing a competing diol compound such as sorbitol, which displaces the glycated haemoglobin from the resin.

A key characteristic of this chemistry is its specificity for the glycated cis-diol structure on the sugar moiety rather than for any particular site of glycation on the haemoglobin molecule. Unlike ion-exchange high-performance liquid chromatography (HPLC), which separates haemoglobin variants by charge, boronate affinity methods capture all glycated haemoglobin species — including those glycated at sites other than the N-terminal valine. Because the method therefore measures total glycated haemoglobin rather than specifically the N-terminal glycated fraction defined by the IFCC reference method, careful calibration against IFCC-aligned reference standards is essential to ensure results are comparable across platforms. The reversibility of the boronate–diol bond under controlled conditions makes it well suited to automated laboratory workflows.

How Boronate Affinity Methods Measure HbA1c in Practice

A haemolysed blood sample is passed through a phenylboronic acid resin; glycated haemoglobin binds, is washed, then eluted and quantified by absorbance at 415 nm to calculate the HbA1c ratio in mmol/mol.

In a typical boronate affinity assay, a haemolysed blood sample is introduced onto a column or cartridge packed with phenylboronic acid-functionalised resin. The assay proceeds through several defined steps:

• Sample preparation: Whole blood is haemolysed to release haemoglobin from red blood cells.

• Binding phase: The haemolysate is passed through the boronate resin at the appropriate pH (usually 8.0–8.5), allowing glycated haemoglobin to bind selectively.

• Wash step: Non-glycated haemoglobin and other proteins are washed away, reducing background interference.

• Elution: Glycated haemoglobin is released using an acidic buffer or a competing diol reagent.

• Detection: Both the glycated and total haemoglobin fractions are quantified, typically by absorbance at 415 nm (the Soret band), and the HbA1c value is calculated from their ratio and expressed in mmol/mol.

Boronate affinity technology is employed in several point-of-care (POC) devices as well as laboratory-based analysers. One example of a platform incorporating boronate affinity principles is the Trinity Biotech Premier Hb9210; other manufacturers offer devices based on alternative analytical principles such as ion-exchange HPLC or immunoassay. Clinicians and laboratory staff should verify the analytical principle of any specific platform via the manufacturer's instructions for use or the NGSP (National Glycohaemoglobin Standardisation Program) method certification listings.

POC devices using boronate affinity can deliver results within minutes from a fingerprick or venous sample, supporting timely clinical decision-making during a consultation. However, an important UK practice point is that HbA1c for diagnostic purposes should be measured using an accredited laboratory method with demonstrated traceability to the IFCC reference system. POC HbA1c results should not generally be used to diagnose diabetes unless the device and the process have been validated and operate within a robust POCT governance framework compliant with ISO 22870 and ISO 15189, as required by UKAS-accredited services. POC testing is most appropriately used for monitoring glycaemic control in people with an established diagnosis.

Accuracy, Interference, and IFCC Standardisation

Boronate affinity methods must be traceable to the IFCC reference system and are generally less susceptible to haemoglobin variant interference than ion-exchange HPLC, though altered red cell lifespan and haemoglobinopathies can still affect results.

The accuracy of any HbA1c method depends on its alignment with internationally recognised reference standards. The International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) has established a definitive reference method based on mass spectrometry and capillary electrophoresis, which specifically measures the glycated N-terminal hexapeptide of the haemoglobin beta-chain. All methods used in UK NHS laboratories — including boronate affinity platforms — must be traceable to the IFCC reference system, with results reported in mmol/mol as mandated since 2011. Some laboratories also provide the DCCT-aligned percentage value alongside the mmol/mol result for continuity, though mmol/mol is the primary UK reporting unit.

Boronate affinity methods are generally considered robust, but several sources of interference warrant clinical awareness:

• Haemoglobin variants: Conditions such as sickle cell trait (HbS), HbC, or HbE may affect results on some platforms. Boronate affinity methods are often less susceptible to interference from common variants than ion-exchange HPLC, but laboratories should flag samples from patients with known haemoglobinopathies. Elevated fetal haemoglobin (HbF) may also affect some methods; clinicians should consult their local laboratory for method-specific interference information.

• Altered red cell lifespan: Haemolytic anaemia, iron deficiency anaemia, or recent blood transfusion can falsely lower or raise HbA1c by altering the proportion of older, more glycated red cells. In these situations, HbA1c may not accurately reflect average glycaemia, and alternative markers such as fructosamine may be considered.

• Carbamylated haemoglobin: In advanced renal failure, carbamylation of haemoglobin can interfere with some ion-exchange HPLC methods by altering the charge-based separation of haemoglobin fractions. Carbamylated haemoglobin does not contain cis-diol groups and is therefore not expected to interfere with boronate affinity assays, which is one advantage of this analytical approach in patients with chronic kidney disease.

• Other matrix effects: Severe hypertriglyceridaemia or hyperbilirubinaemia may affect some platforms. Labile (pre-ketoamine) intermediates are generally removed during sample preparation or the wash step in boronate affinity methods, reducing their impact on results.

Clinicians should interpret HbA1c results in the context of the patient's full clinical picture, particularly when haematological conditions are present. Participation in external quality assurance schemes such as UK NEQAS for HbA1c is required under UKAS ISO 15189 accreditation and is strongly recommended by professional bodies, ensuring ongoing method performance and comparability across laboratories.

Clinical Use of HbA1c Testing in NHS Diabetes Care

NICE recommends HbA1c monitoring every three to six months for people with type 2 diabetes, with individual targets of 48–53 mmol/mol depending on therapy and hypoglycaemia risk; accredited laboratory methods are required for diagnosis.

Within the NHS, HbA1c testing underpins the entire pathway of diabetes diagnosis, monitoring, and treatment optimisation. NICE guidelines (NG28) recommend that people with type 2 diabetes have their HbA1c measured every three to six months until stable on treatment, then every six months thereafter. Individual HbA1c targets are agreed between the patient and their clinician, typically aiming for 48 mmol/mol (6.5%) in newly diagnosed type 2 diabetes managed by lifestyle or metformin alone, or 53 mmol/mol (7.0%) where additional therapies are in use or hypoglycaemia risk is a concern.

For people with type 1 diabetes, NICE (NG17) recommends a target of 48 mmol/mol where achievable without problematic hypoglycaemia, acknowledging that individual circumstances vary considerably. Continuous glucose monitoring (CGM) is increasingly used alongside HbA1c in type 1 diabetes management, providing complementary information about glucose variability that HbA1c alone cannot capture.

As noted above, HbA1c is not suitable for diagnosis in children and young people, during pregnancy, where type 1 diabetes or rapidly evolving diabetes is suspected, or where conditions affecting red cell turnover are present. In these circumstances, plasma glucose-based testing should be used. For monitoring purposes, boronate affinity-based POC devices have an important role in community and primary care settings, where same-appointment results can facilitate more responsive clinical decisions — for example, adjusting medication or reinforcing lifestyle advice during a single consultation. For diagnostic purposes, an accredited laboratory method with IFCC traceability should be used; POC devices should only be used for diagnosis where they have been formally validated within a POCT governance framework meeting ISO 22870 and ISO 15189 requirements.

NHS England's Quality and Outcomes Framework (QOF) includes HbA1c-based indicators as part of diabetes care quality metrics, incentivising regular, standardised testing in primary care.

Patients should be advised that a single HbA1c result represents an average and does not capture day-to-day glucose fluctuations. If a patient experiences symptoms suggestive of hypoglycaemia, significant weight loss, or unexplained deterioration in wellbeing, they should contact their GP or diabetes care team promptly, regardless of their most recent HbA1c value. Regular testing, interpreted alongside clinical assessment, remains the cornerstone of safe and effective diabetes management in the UK.

Frequently Asked Questions