HbA1c UNCOVERED: The Science of Boronate Affinity Chromatography

This article walks through what HbA1c actually is, all the way to how an analyser works at the molecular level, with a focus on the Boronate Affinity principle — an approach with clear advantages in the Thai population.

Part 1 · The Basics

What is HbA1c?

Red blood cells contain a protein called haemoglobin, which carries oxygen. As glucose circulates in the bloodstream, it gradually attaches itself to haemoglobin without needing any helper molecule — and once attached, it rarely comes off. HbA1c is therefore not “the blood sugar level” but the proportion of haemoglobin that has sugar attached to it, relative to total haemoglobin, reported as a percentage.

A simple comparison: a fasting blood glucose test is a photograph of that single moment, while HbA1c is a logbook that has been recording glucose levels over the past 2–3 months.

The reason it reaches back 2–3 months is that red blood cells live roughly 120 days. Once sugar attaches, it stays attached until that cell reaches the end of its life. The measured value is therefore a weighted average, with roughly half of it coming from the final month before the blood draw. And because it is a cumulative value, HbA1c does not swing with meals or stress, and no fasting is required before testing.

Part 2 · The Boronate Affinity Principle, Visualised

The core idea in one sentence: this method separates by the structure of the attached sugar, not by the electrical charge or shape of the haemoglobin protein itself.

This distinction is the key to why the method tolerates abnormal haemoglobins so well. If the analyser looks at the protein, it gets confused when the protein changes shape. But if it looks at the sugar, then no matter what the protein looks like, sugar is still sugar.

1. Sugar attaches to haemoglobin — this happens inside the patient, not inside the analyser. The important point is that after attachment, the sugar molecule still carries two hydroxyl (–OH) groups sitting next to each other and facing the same direction. This structure is called a cis-diol, and it is the “key” the analyser goes after.
2. The capture site locks onto the paired –OH groups — inside the analyser is a membrane or resin coated with boronic acid. The boron atom must bind both –OH positions simultaneously to hold on — like a two-pin plug that only locks when both pins are inserted. Haemoglobin carrying sugar is therefore captured, while anything without a cis-diol flows through.
3. Count the ratio, then report as a percentage — the analyser quantifies both fractions from the same sample, calculates the ratio, then aligns it to the NGSP and IFCC international standards before reporting a percentage that is comparable with results from other laboratories.

Part 3 · Why Abnormal Haemoglobins Do Not Interfere

• A variant is a change in the protein — HbS changes the amino acid at position 6 of the beta chain, HbE at position 26. This alters the charge and shape, so charge-based methods lose their reference point.
• But boronic acid does not bind to the protein — it binds only to the cis-diol, a structure found exclusively on the sugar molecule.
• The consequence — changing an amino acid at any position neither creates nor removes a cis-diol. The amount of attached sugar is unchanged, so the analyser counts the same value.
• The same reasoning also excludes carbamylated and acetylated Hb — for the identical reason: they have no cis-diol.

Part 4 · Comparison with Other HbA1c Methods

No single method wins on every front. Ion-exchange HPLC provides the most information, because the chromatogram is visible and flags the presence of an abnormal variant. Boronate affinity has the advantage of tolerating interference from both variants and chemically modified haemoglobin derivatives, without requiring a complex environmental control system.

Part 5 · Why This Matters Particularly in Thailand

Of every 100 Thai people, roughly 30–40 carry thalassaemia trait or some form of abnormal haemoglobin. The most common type is HbE, found at especially high rates in the northeastern region.

Most of these individuals do not even know they are carriers, because they are healthy, not anaemic, and have normal red cell lifespan — and they develop diabetes just like anyone else, so they need routine HbA1c monitoring.

The point to watch is that with a charge-based analyser, the result can be off without anything flagging it. The number printed out looks perfectly normal, when in fact the value may not be accurate.

In patients with kidney disease and those taking high-dose aspirin, another substance — not sugar — attaches to haemoglobin. These substances change the protein’s charge in a way that resembles what happens when sugar attaches, so a charge-based analyser may mistake them for sugar and count them in.

The boronate principle, by contrast, does not care what charge the protein carries. It looks only for paired –OH groups. These two substances have no paired –OH groups, so they never bind and are never counted.

Part 6 · Limitations to Know Before Interpreting a Result

This section matters as much as the strengths. Failing to address the limitations is one of the leading causes of misinterpretation.

• Shortened red cell lifespan gives a falsely low value — for example, clinically significant thalassaemia (homozygous HbE, HbE/beta-thalassaemia, HbSS, HbSC), haemolytic anaemia, acute blood loss, treatment with erythropoiesis-stimulating agents, or splenomegaly.
• HbF above roughly 10–15% gives a falsely low value — this affects boronate affinity, immunoassay and enzymatic methods alike. Published data for the boronate affinity method indicate no interference below approximately 10.7% HbF.
• Untreated iron deficiency gives a falsely high value — the value falls on its own once iron is replaced.
• Pregnancy shifts the value in both directions — lower in early to mid pregnancy, and possibly higher in the third trimester if iron deficiency is present. The ADA therefore does not recommend HbA1c as the primary criterion for diagnosis during pregnancy.
• It cannot identify the variant type, because it does not resolve individual haemoglobin peaks. It is therefore not a substitute for thalassaemia screening.
• End-stage renal disease and dialysis patients — although carbamylated Hb causes no interference, red cell lifespan is usually shortened, so glycaemic control is still underestimated. Glycated albumin or direct glucose monitoring is preferable.

Part 7 · Practical Guidance and Conclusion

Boronate Affinity HbA1c Analysers from NS Promedica

NS Promedica supplies point-of-care HbA1c analysers built on the Boronate Affinity principle described in this article. They are NGSP-certified and traceable to the IFCC system, require only a small blood volume, deliver results within minutes, and come at an accessible price.

A1C Chek Express – HbA1c analyzer (Boronate Affinity Chromatography) – NS Promedica

A1C Chek Pro – HbA1c analyzer (Boronate Affinity Chromatography) – NS Promedica