In the Lasers 4 Life project, physicist Marinus Huber, a doctoral student, examines the blood samples with infrared laser light in the LEX Photonics Laser Lab at LMU Munich. After the light has passed through the sample, Huber once again analyses the spectrum of the transmitted light, which differs from that of the incident beam. This is because the molecules present in the blood have interacted with, and absorbed some of the incident light at specific wavelengths. Since the precise molecular composition of blood varies from one individual to the next, the pattern of light absorption serves as a “chemical fingerprint” of the sample donor’s metabolic state.

The next question, of course, is whether or not one can distinguish healthy individuals from people with cancer based on the laser spectroscopic analysis of blood samples. In their experiments so far, the L4L team has shown that the new laser technology is 50 times more sensitive than conventional analytical methods. In the next step, Marinus Huber will ask whether this increase in detection sensitivity is sufficient to allow him to reliably detect the presence of cancer cells.

Trustworthy medical diagnoses depend on the sensitivity, discriminatory power and reproducibility of the tests employed. Ensuring that these criteria are met is a crucial component of the Lasers4Life project (L4L). A blood test based on the application of laser spectroscopy needs to be extremely sensitive and, among other things, this means that the samples used must be prepared in a very particular way.

In the L4L team this task is in the hands of Cristina Leonardo, a chemist. She is developing a procedure which allows one to cleanly divide the soluble (non-cellular) portion of the blood sample into a protein fraction and a metabolite fraction. Both fractions are made up of organic substances that are produced in cells, and released into the circulation. Most importantly, their detailed composition is expected to differ in subtle ways, depending on whether or not cancer cells are present in the donor. The L4L team will use laser light to characterize the molecules in the two fractions, thus providing a “molecular fingerprint” for each donor. The ultimate aim is to identify the specific features of the fingerprint that reliably indicate the presence of cancer cells in the body.

We, the L4L team, would like to take this opportunity to express our heartfelt thanks for the generosity and readiness to donate on Thursday and Friday last week at the LMU Frauenklink. Now it's time to evaluate the samples.

An Important visitor made an appearance at the Center for Advanced Laser Applications and the Laboratory for Extreme Photonics last Friday. Dr. László Palkovics, Minister for Innovation and Technology was on the research campus in Garching and visited the two laser research facilities at the Ludwig-Maximilians-Universität.

While there, he received a tour of the large laser systems in the laboratories from Professor Ferenc Krausz and Dr. Andreas Döpp. The minister was particularly impressed by the enormous developments that laser technology has made in recent years and the associated opportunities for their use in medicine. Of particular interest for him, was the BIRD project and its blood analysis using laser light. Collaboration with clinics in Hungary in the framework of the project is currently being planned.

Immediately after taking the blood, the L4L team processes your donation in the biobank. Before the blood can be examined under laser light, several careful processing steps are necessary.

About half of the blood is fluid: the blood plasma. It is in this fluid that the blood cells can be found. In just one milliliter, there are around five billion red blood cells, 200 million platelets and five to ten million white blood cells. Our study assistants eliminate the solid components of the blood, so that only the liquid is retained. This happens via a kind of spin process during so-called centrifuging.

Since the blood is not immediately examined under laser light, it is first transferred to small cannulas and then stored in special refrigerators at minus 80 degrees Celsius. This slows down all biological processes to such an extent that it is possible to work with the valuable samples for up to several years.

Junior researchers at LMU Munich are also actively involved in the Lasers4Life project. One of them is Maša Bozič. As part of her Master’s project, she is using visible light to analyse blood samples, before they are examined with the newly developed near-infrared laser.

Both of these approaches use spectroscopy to characterize blood serum and blood plasma. The term ‘serum’ refers to the liquid phase obtained after whole blood has been allowed to coagulate, which therefore contains no clotting factors. Plasma retains all the normal clotting factors, but activation of the cascade of enzyme reactions that leads to blood clotting is prevented by the addition of an anticoagulant. Coagulation is normally triggered by damage to blood vessels. This in turn causes blood platelets to adhere the damaged vessel wall and ultimately leads to conversion of the protein fibrinogen into the fibrin network, which forms the mature clot and seals the wound.

Both serum and plasma are obtained from whole blood by centrifugation in the presence (plasma) or absence (serum) of an anticoagulant. The centrifugation step serves to remove the red and white blood cells (together with the clot, in the case of serum). Maša then subjects both samples to optical spectroscopy with visible light. As the beam passes through the sample, certain wavelengths of the incident light are absorbed by the substances present in the solution. The changes observed in the transmitted spectrum therefore provide information on the composition of the non-cellular fraction of the blood.

In this way, one can determine the concentration of certain proteins and lipids based on the characteristic pattern of absorption of the incident light. The method therefore allows Maša to establish the extent of day-to-day variation between samples taken from the same individual, or the range of variation between different individuals. This provides a baseline that allows one to assess whether differences in the absorption spectra lie within the normal limits of variation, or are indicative of pathological changes that reflect the presence of disease. The results also provide initial insights into the scale of the differences between the spectra obtained from cancer patients and control subjects.

Maša Bozic’s work therefore yields an essential reference for subsequent spectroscopic analyses with infrared laser light in the L4L project, as both spectroscopic methods rely on the same principle of selective absorption. However, infrared spectroscopy is far more sensitive than conventional spectroscopy with visible light. It therefore provides far more comprehensive and detailed information on the diversity of substances present in the samples, and should allow one to identify those that may be linked to the presence of malignant cells in the blood donor.

Nathalie Nagl has been awarded a doctoral scholarship by the Bischöfliche Studienförderung Cusanuswerk. She has already written her master’s thesis in Dr. Oleg Pronin’s group in the LAP team and can now continue her work as a doctoral student. Nathalie is working on a new, pulsed laser light source that emits near-infrared radiation. It uses a Cr:ZnSe crystal as a laser medium, as well as novel diodes, which are needed to pump the crystal.

The system is designed to detect specific molecules in biological samples. The molecules that researchers are interested in are often very weakly concentrated and thus difficult to find. For this reason, the laser source used must produce as little noise as possible and send out extreme strong light at very specific frequencies. Molecules each react only to a well-defined frequency of light.

Nathalie now wants to push the laser deeper into the infrared range. This could make it possible to detect an even wider range of molecules

At the beginning of October Dr. Frank Fleischmann joined the Broadband Infrared Diagnostics (BIRD) team led by Dr. Mihaela Zigman. Fleischmann began his career in biology as a botanist, but later switched to medical research. Before taking up his present position as a member of the BIRD team, he worked for a commercial provider of genetic tests, including the genotyping of cancer patients, for example.

Fleischmann’s role in the BIRD team is akin to that of an archivist. He is responsible for the cataloging and storage of blood samples. Needless to say, accurate documentation and painstaking handling of test samples are of fundamental importance in medical research. After all, its ultimate goal is to produce a therapeutic agent or procedure that will be used to treat real patients every day. Fleischmann is also in charge of the database specially developed for the Lasers4Life project, and meticulously documents everything done with each and every one of the vital samples in his care.

At the moment, the samples of blood plasma and the sera obtained from them are being stored at a temperature of −80°C. However, even this temperature is not low enough for long-term storage of such samples, as slow ice recrystallization alters their consistency, and after a certain time they have to be discarded. Fleischmann is working on an automated cooling system based on liquid nitrogen as the refrigerant, which will allow the samples to be kept at temperatures as low as −180°C. This is sufficiently cold to inhibit ice recrystallization in the liquid – and under these conditions, the constituents of the various blood fractions will remain unchanged for decades. Thus, as even more advanced methods of laser spectroscopy are developed in the future, the new system will enable the BIRD team to re-examine the samples already collected.

Imagine going to your GP for a screening test. Within minutes the test tells you whether or not you will get cancer in the near future. Would you really want to have access to such information? The fact is, it is already there in our bodies — you just have to decode it. This is the task of Dr. Kosmas Kepesidis, a physicist and data scientist who has recently joined the Broadband Infrared Diagnostics (BIRD) team at the Laboratory for Attosecond Physics. His name is fitting — ‘Kosmas’ derives from the Greek for ‘cosmos’ or ‘world’ and that is exactly what Kosmas studies: the microcosm of molecules in our blood. He does this with the help of algorithms, in other words, numbers.

The scientists on the BIRD team are developing a medical diagnostics tool to detect cancer based on the analysis of infrared light waves. These are emitted when ultrashort laser pulses excite molecules in the blood. The resulting spectra contain fingerprint-like information about the blood’s molecular make-up and thus the state of the patient’s health.

The problem is that, unlike the abstract models used in physics, biological systems are highly complex. Thousands of data points are collected and no one quite knows what to look for. Who even has the time to sift through them? Kosmas is therefore developing software which uses machine learning algorithms to carry out predictive modelling. In other words, he uses advanced computational methods to predict outcomes, such as whether a given molecular fingerprint is an indicator of early-onset cancer.

First, thousands of samples are collected from patients with and without cancer. Thus, we end up with two massive mounds of data. Eventually, the goal is to create further stacks to differentiate between distinct types of cancer. These mounds of data are then pre-processed. For instance, decisions have to be made about which patterns constitute ‘noise’ and can be ignored. Next, Kosmas performs a so-called ‘dimensionality reduction’, i.e. he ‘zooms in’ on those features of the data that are relevant. The third stage is the search for a model: which algorithm is most suited for cancer diagnostics? Kosmas hopes to use artificial ‘neural networks’, algorithms which very roughly approximate biological nervous systems in how they process high-level, as opposed to low-level information. Such algorithms work with abstract patterns rather than zeros and ones like conventional computer programs. For this to work, Kosmas needs lots of data, which are currently being collected in hospitals around the world. Finally, once a model is found, Kosmas will expose it to rigorous testing.

He hopes that his research will culminate in a simple-to-use app that enables physicians to analyse blood samples on the spot. How long it will take to develop this software is as yet uncertain. ‘There are a lot of factors involved.’ And while a screening test which, having analysed a blood sample, outputs either ‘cancerous’ or ‘non-cancerous’ is clearly sufficient for daily life at the doctor’s office, scientists of course want to know exactly which features of the molecular fingerprint are responsible for such diagnoses. But Kosmas relishes the risks and uncertainty involved in doing cutting-edge science. The tools change constantly. ‘I do not know what my work will look like in a few months’ time. I expect it to change a lot’.

Our Broadband infrared molecular fingerprinting project is expanding its network. Joint work with Hungarian clinical centers has started. We would like to welcome Dr. Gábor Csík, a specialist in managing clinical trials, who just joined the team of Mihaela Zigman. Gábor will be in charge of building up and heading the L4L network of medical collaborations in the region. Joint work with the Szeged Medical University, the Koranyi National Institute of TB and Pulmonology Budapest and the Heart and Vascular Center of Semmelweiss University in Budapest has already begun. The plan is to expand the existing clinical network for infrared molecular fingerprinting to tackle disease detection across Hungary.