Vega lmu

Flower morphology was observed under an STM 800 stereomicroscope (mikroLAB). Photographs were taken using a Nikon Coolpix 4500 camera (Nikon, Tokyo, Japan).
Fresh pollen from anthers of both species was collected and glycerin–gelatin preparations with the addition of basic fuchsin were made [55 (link)]. Basic fuchsin stains the exine and the intine. Photographic documentation was made with the use of a Nikon Eclipse 400 light microscope (Nikon, Tokyo, Japan) coupled to an Olympus DP23-CU camera (Olympus, Tokyo, Japan) working with CS-EN-V3 cellSens Entry V3 software.
Tilia pollen grains were also observed using a scanning electron microscope (SEM). Aerobiological samples were collected in 2020 using Durham devices located at a height of 2 m in the Botanical Garden of Maria Curie-Skłodowska University in Lublin. Slides covered with double-sided adhesive tape were placed in the devices. Fragments of the tape with biological aerosol samples were coated with gold using an Emitech SC 7640 sputter coater (Polaron, Newhaven, East Sussex, UK) without application of standard preparatory procedures used in scanning electron microscopy. The preparations were observed and photographed in a TESCAN/VEGA LMU (Tescan, Brno, Czech Republic) scanning electron microscope equipped with a TESCAN attachment for digital processing of microscopic images.

The nectaries (n = 10) for the analyses were collected from different flowers and plants. The samples were fixed in 4% glutaraldehyde in 0.1 M sodium phosphate buffer (pH 7.0) for 12 h at room temperature. Next, they were washed in the same buffer four times at 20 min intervals and dehydrated in increasing concentrations of an acetone series (30, 50, 70, 90, 95%). The dehydrated samples were critical-point dried in liquid CO₂ using Bal-Tec CPD 030 (Bal-Tec, Balzers, Liechtenstein). The plant material prepared in this way was mounted onto stubs and gold sputter coated (thickness approx. 10 μm) using a Emitech SC 7640 sputter coater (Polaron, Newhaven, East Sussex, UK). The specimens were analysed under a TESCAN/VEGA LMU (Tescan, Brno, Czech Republic) scanning electron microscope at an accelerating voltage of 30 kV.

Fragments of the corolla petals and calyx as well as stamens were fixed in a 4% glutaraldehyde solution in 0.1 M phosphate buffer (pH 7.0). After 12 h incubation at a temperature of 4 °C, the samples were washed in the same buffer four times at 20 min intervals. An ethanol series (30, 50, 70, 90, and 95%) was used to dehydrate the plant material. Next, the samples were immersed three times in absolute alcohol and transferred to acetone. The samples were critical point dried in liquid CO₂ using Bal-Tec CPD 030 (Balzers, Liechtenstein).
Dried fragments of the examined floral parts were glued onto stubs with the use of a double-sided carbon tape. The samples were coated with a 10 μm gold layer using a Polaron SC 7640 sputter coater (Emitech). A scanning electron microscope TESCAN/VEGA LMU (Tescan) at an accelerating voltage of 30 kV was used for the examination of the material.